Synthetic, plasma-free, transfusible storage medium for red blood cells and platelets

1326~ SYN~HETIC, PLASMA-F~EE, ~ FUgI8LE STORAGE MEPIUM FOR REI~ BI.OOD C~l,LS AND PLATELE~S BACXGROUNr) OF THF INV~NTION 1.

Tecl~nical e~lelB The pre6en~ ~nventlon relate~ to ~ 6ynth~tic, pl~m~-~ree ~u6pen~Lon med~um for red blood cell~ ~nd pl~tel~ts. More partlcularly, the pre~ent ln~ention rel~te~ to a ~ynthatic preservatlon medlum for red blood cells or plat~let~ whlch (1) 1~ free of blood pla6m~ ~nd other protelns, ~2) extend~ red blood celi or pl~tele~ ~helf 11~8 whlle malntalning the quallty of concentrat~s of the6e component~ durlng 6tor~ge for tr~n6fu~ion and (3) 18 free of une~oentl~l organlc compound~.

. State of the Art 8100d ~8 compo6ed of two major portlons. The6e portlons can be recognl~ed ~hen ~ 6p~L~n of blood 1 dr~wn an~ clotting 18 prevented. ~h~t portion of the blood whlch settles to th~ botto~ o~ the v~fi~el holdlng the 6peclmen 1~ termed the "formed el~entr~." The formed element~ comprlse red blood ~ell6 and other partlculate component~ 6Uch A~ whlte blood cell8, red blood cells, ~nd pl~telet8. Pl~telet6 ~re also known throm~ocyte~. ~he formed elemQnt~ arH _ _ ~ ~ J 1326~140 ch~racterlstlcally 40 to 50 percen~ o~ the bulk ot normAl hum~n blood. The cloudy llquld whlch doo6 not dettl~ ln ~ blood ~poclmen 1~ the portlon o~ the ~looa known as pl~sma. Pl~m~ i8 prlmarlly water, but cont~ln~ lnorganlc and organ~c su~stancefi a~ well ~ dls601ved gase~ and mlscellanoous ~orel~n ~ub~t~nc~6.

~h~ lnorganlc ~u~st~nce~ contaln~d ln ~lood pl~ma ~e prlmarlly Qlcctrolytos. ~he mo~t 41gnlflcent of the~ olectrolyte~ aro prosonted ln T~ble 1 in ~h~ concentratlons typ~cally found ~n he~lthy, hu~n blood plA~ AB~E~.l ~dlum 142.0 mEq/l PotA441u~ 4.3 m~q~l Calclum 5.0 m~q~ Magne~lum 3,4 ~ Chl~rld~ 10~.0 m~q~l ~lcar~onato 27.0 m~q~l Pho~ph~te 2.3 m~q~l 8ul~ato 0.6 ~q~l ~ ~2~ The moBt ~i~ni~lcant o~g~ni~ SubBt~nceB ~ound ln the pln~m~ ~re lactlc ncld, ur~, a~lno acld~, crcAtlnlne, gluco~e, hormone~, proteins, Albumlnr, ~nd globullns, Modern medlcin~ h~8 b~e~ developlng 801utlon~ that ~re addQd to blood in vlvo And/or mlxsd wlth blood ln vltro~ Produ~t~ th~t ~re used for adding to blood ~n vlv~ ~e prlm~rlly u~ed for lntr~veneou6 feedlng, phar~acoutlcal ~ehlclc6, ~nd/o~ ctrolyte replAcement ln patlente who are b~dfa6t. The4e solution~ are pr~arll~ comprl6~d of water that contains dextro~e and, opt~onally, olectrol~e~. Dextro~e ~6 typlc~lly pre6ent ln t~e6e solutlon6 ln a~out a 5 pe~çent conoentratlon and pro~ldeB ~ nutrlen~ for blood çellc or ti6suo cells.

~he ele~trolytc~ conta~ ned ln these ~olutlon~ vary wl~ely, The ~ol~tlon~ that contaln el~c~rolytos that mo6~ 0108ely re6emble blood plasm~ cont~in a plucAl~ty of the electrolyto6 pre~ented in Table 1. A ~peclflc example of a dextro-e and l-ctrolyte 601utlon ~ult~blo ~o~ ln vlvo addlt~on in biood 1~ Lock~-~inp-r'~ 601utlon.

Tho formula fo~ ~Ock~-Ringor~ ~olutlon 1 pro~entod ln ~blo 2.

~ v 1326~0 TA~LE_2 R~g~nt ~odium Chlorlde 9.0 Gm Re~gent Po~a~lum Chlorlde 0.42 Gm Reag~nt c~lcium Chlorldo O . Z4 Gm R~gen~c M~gn~ium Chloelde O . Z Gm Sodlum BlcAr~onat~ O . 5 Gm D~xtross 0 . 5 Gm W~ter, ~c~n~ly dl~t~lled ~rom ~ hArd 511~81i~ flA~k~ ln ~ ~u~rlcl~nt quanti ty, _~ to Dl~lte 1000 ml Other ~olutlonr ~ultAble for the ~ddltlon o~ blood ln vlvo c~n b~ found ln ~emington's Ph~rm~ceutlctll - Scl~nce~, M~ck Publl~hlng Company, 14th ~dltlon ~19'10~, p~ge~ 815 to 847.

801utlon~ th~t are ~dded ~o bloo~ in vltro ~re elth~r slmplc ~ntlco~gul~nt nutrlent mlxturo~ prlnc~p~lly d~gned ~or jr~s~r~lng whole blood or ~epar~tcd compon~nt~ ~or llmlt-d p~rlod6 of tlm~ or ~rspcclrlc nutrl~nt llolutlonl~ optlml2ed ~or prolong~d pro~crv~tion o~ specl~lc blood compon~ne# ~uch ~ rcd c~ , plAt~l-t-, whle~ ~lood coll~, or m~xtur-~ of th~o compon~nt- ~h-n pl~t~lot- ~r~ ~nclud~d ~ on~ co~ponont o~ blood th~t 1~ to b- aollcctcd And pr-o-rv~d --4- 1326~0 ln vltro wlth red blood cells, lt ~ do~r~ble to u~e lo~ ~ctlve ~ntlcoa~ulant.

~ he mo~t ~roqu~ntly u60d antlcoagulant added to collected whole blood ld known a8 "acld-eltrate-dextro~o" or "ACDn. ~hls ~ntl¢o~gul~nt ~olutlon cont~lns ~ mixtur~ of antlco~gulant ~nd nutri~nt ~p~cl~icallys ~1) citrlc acld and Rodlum oltr~te in opti~um concentratlons eu~flclent to chol~t~ oalcium wlthout or~atlng ~n unphy~lologlchl p~ and ~2) dextro6e ln concentr~tion6 ~ufflciont for short ter~ pr~s~rvatlon of blood or component~, o~poclally tho red bloo~ cell~ A le8E ~cldlc solution thAt ~as been found ~irAblo to pre~erve both whole blood and other c~llul~r f~a~tions 1~ known a8 "~ntlaoagul~n~ cltr~to-phosph~t~-dextro~e solutlon~ or "CPDn. ~ho ~o~ponont- of antlcoagulant cltr~te-p~o~phate~dextros~ olutlon are pr-~ented ln Table 3.

f_, ~ ~ 1326~0 TAsLE 3 Cltrlc Ac~d ~nhydrous) 3.0 G~ ~odlum Cltrate ~dlh~drate) 26.3 ~m Sodlum ~lphosph~te ~monohydr~te;

N~H2Po5N2o) 2.z2 am Doxtro~e 25.5 Gm w~er or In~octlon, ln ~u~lc~en~ q~ant~ty to ~ke 1000 ml 8peol~1c elements of the partlculato component c~ blood c~n be ~oparate~ ~nd better pro~er~ed for latoc tran~u#ion at dlfferent temperAture~. S~ple 6ep~r~tlon proco~e~ c~n bo u~ed to c~lloct ~nd pr~r~e wh$te blood cell~ ~nd plAtelet6 whlch a~e be~t 6tored at 22C r~h~r ~han 4C whloh ~ optlon~l ~or red cell6.

8torase of red cel~B ~t ~C ~or transfu610n r~fiult~ ~n detorlor~lon o~ th~ cell6 over ~ perlod of tlme.

~hl~ dctorloratlon of tho c~116 l~ tormod "~torAg~ lc~lon" and 1~ oh~ractorl~ed by ~nor~a~ed h~moly~l- And d~ro~s-d vl~b~l~ty r~ultlng ln loworod po~t-trans~u~lon ~ur~l~al o~ th~ r~d blood c~ . Tho ~lt~r~t~ons a~soc~od wlth rtorAge l-~lon lnclud~ lntr~collulAr chango~ ~uch A~ decre~s-d pota~41u~ lon and ~cre~od ~odlum An~ ~lc~um lon concentratlon~, tho 132~ 10B~ of 2,3-diphospho~lycerate (2,3 DPG) re6ultlng ln altered oxygen-tr~nsfer char~cterl~tloe, the deplotlon of adono~ine ~Fiphoephate ~A~ and ~embrane ~keloton alteratlons resultlng ln deceee~od doo~m~bllity and ~ncre~ed fraglllty ~ ed cell6 were pr~viou61y, typlc~lly ~tored ln a comblQed nutrlent~Antlcoagulant mlxture consl~ting o~ 1 pArt nutrient~anticoagulAnt to 7 parts pla6ma (acld~cltr~te-dextro~s) The progreeslon o~ ~torage le~ion wa~ the ll~itlng factor to the ~helf life o~ ~tored red cell~ ~he shelf llfe o~ ~tored red b~ood cell~ wA~ geneF~lly l~it~d to 21 days~ More recently, lmproved red cell vl~lllty wlth ~orago perlod~ up to 4g day~ ha~ been ach~eved w~th ~d~l~lve~ ~pecl~lc~lly mlxed wlth rsd cell~ ~ftor the pl~m~ and pl~t-let~ hav~ been romoved ~yp~cally, the~o addltlve ~olutlons aro a 41mplc ~allne ~olutlon fortifled wlth ad-nln-, gluco4e, ~nd other, mlnor con~tltuent~ that are a~dod to ret~r~ developm-nt of h~oly~l6 and lor~ of 2,3 DP~ The ptl~ry contalner ur~d wlth th-r- addltlv- ~olutlons contaln- only a ~mplo ant~coa~ulant whlch foe oonv~nl-nc- ha- b~n CPO a~ do~crlbod pr-vlouely ~t~r ooll~ctlon, thantlco~gulato~ whol- blood 1~ centrl~uq~d and thplat~l-t~rlch pla~ma and eed blood c-ll- ar- tean6~rod ~326440 to a 0~tclllto b~g. ~ho rod c~ll ad~ltl~e solutlon 1 thon mlxod wlth tho r~d c0116.

Mlxlng ~h~ ltlvo eolutlon lnto tho sop~ra~od ro~ blood c~ provent~ con~atnlnAtlon o oth~r s~p~roltc~ co~ponent~ ~ tho blood wlth th~ ro~ cell ~ddltive ~olut~on.

De~lr~bla ~low p~op~rti~ ¢An b~ malntaln~d or the ro~ c~ by u~lng ~n app~oprl~to ~olu~o o~ tho ~ddltlrc ~olutlon.

Nu~morou6 additlr~ solutlons tor red blood ooll~ exl~t alther ~18 co~m~rclal or re~o~rch product~..

Shc typlc~l ~dd~t~ve solutlon ~or re~ blood 00116 ~8 d~rlv~d ~eom ~ no-~d~n~n~-g~uco~ (8AG) ~d~u~. 8A0 d~volopod ln 8wodon ln tho ~ld-~g70~s an~ provldo~ acco~t~b~ oryth~ooytc ~rrlYal ~tar 35 day~ of ~tor~go rathor th~n ~ho 21 dayr that WaB prer~ou~ly ~vall~blo wlth co~bln~ antl-co~gulan~ nutrient 801~tlon~. 6A~ ~alntaln~ ~co~ptable AT~ lev~l~ throughout 3S days o~ ~torag~. ~h~ 6torag~ o r~ blood cell~ for 35 day~ ln 8Aa ~o~ult~ ln ~n avora~ h~oly~l~ o~ pproxl~toly 1 p~roont by th~ lart day'o~ torag~ whlch ~a~ ~uc~ gr~ator tban th~ 0.1 p~r~nt nor~lly ~oalat~d wlth ~Ith-r CPD Or ACD. $h~ lncr~a~-d h-~oly~l~ o~ r~d ~lood c~ ln B~G, wh~n co~par~d to 4tor~g- ln ~ ~olutlon o ~o~lu~ oltrat~, d~xteo~, cltrlo ~ald, ~onobarlc 60d~U~ pho!phat-, whlch 14 now ~ortl~l-d wlth ad~nlno (CPDA-l) tor prolongod ~tor~ 35 ~ay~ - `~ ~J iJ 13264q~ b~lle~d to re~ul~ ~rom the aatlon of elthcr whlt- oell protea~es upon the erythroayte membr~ne ln the ~bs~nce o pl~ma protein~ or a~ a fe~ult 08mot1 ~ de~tabllleatlon due to unphyoiolog~c~l ~torAge 601utlon~ .

A modifled ~AG contains mannltol (29 mM) and 18 known b~ the 4~mbol ~AG-M, ~h~ ~dditive solutlon re~ult6 ln membr~ne ~tablll~t~on ~nd d~crea~ed hemoly81q, A produot h~lng ~ ~imil~r formul~ to ~AG-M is tha ~DSo ~ br~nd of storaqe med~um. ADSO ~ whl~h 1 llcensed for commerclol U8e ln the united st~te~ ~nd cont~ins 50 p-rcent ~ore adenln~ ~nd lSO percent more ~luco4e ln ~dd~tlon to 750 mg/dl o2 m~nnltol than doe~ ~G-M. ~hl4 produet 16 produ~od by BAxtor, Inc., 1425 ~ak~ Cook Robd, D~r~leld, Ill~noi~ 60015. The lllgred~entB Or the AD8~ ~rand o~ ~torage ~ dlu~ are preaontod ln T~ble 4.

~h~E 4 D~xtro~ 2.2g 80dlum Chlorld- soo.omg nnltol 750.0mg ~d~nin~ 27.0mg Wat-r ln a ~u~lclont qu~ntlty to m~o lOO.O~l _g_ ~, .... . . .. .

- ' - v 132fi4~ ~he preeence of mannltol ln the ADS0 ~ br~nd ~tor~g~ medlum reduce~ be~olysl~ even t n the p~o~nce of leukooyte pro~ea~e~. The hlgh concentrat~on of d-xtro~c ln th~ ADS0 ~ brand ~torage m~dium 1~ bolleved to have do61 rAble R~eCtB on other red blood cell pa~metersThe ADSo ~ brand 6to~age medlum can of~ectlvely 4tore red cells ~or 4~ days.

However, th~re ~r~ dlsadvAnt~ge6 ~nd problem~ wlth the ouerent gener~tlon of red blood coll storage medi~, These mod~a ~re ionically unbalanced wlth sod~um a~ the only cat~on. Slnce tho ~ctlve cation tran~port sy6tem o~ red blood c~ iB lnhiblted ~t low temperature8, ~ 10B~ ~ th~ norm~l conc~ntration gr~dl~nt occurs ln t~c red blood c~ duo to p~ v- dlffu6ion o~ the c~tlon~.

Thl~ oan OaU6e ~lllng o~ the cell ~nd ev-ntual ly~ The add~tlon to th~e rolutlon~ of ~ mambrane-st~bl~lzer or "o~motlc ~gent~ ~uch a- mann~tol or ~orbltol c*n not ~ully compon~te for the nvlronm~ntal p~rt~rb~tlon o~ the rod c0116 ln the~- ~oiutlons.

Tho p~o~ont gonoratlon o~ rod blood c~ ddlt~vo solutlon~ h~ve A ll~lt-d bu~or~ng capacl~y. Tho~llmltod bu~for~ng ~y~tem~ ~llow the pN o~ tho ~od ~lood ooll concontrAt-- to d~allno to lovel~ bolow 6.5 a~tor about 42 d~yr o~ ~torag~ ~t 4C. At thl~ pH l~v-l and bolow, glycoy~l~ wlth ~TP r~ductlon cau8e8 th~ los4 o~ --10- 1326440 ~od blood cell vlabll~ty during ~torago. The 2, ~ DP~ level 1B al60 m~rkedly ~en61tlve to ~ deçllne ln pH durlng ator~g~. ~o~ of 2,3 DPG re~ult~ ln alt-red red bl~d eell oxygen blndlng chPracterlst~c~.

~ he ~torage of platelet~ ~ n speolal additlve tulutlon~ i~ commonly performed ln 8pcclal or Hsecond generation" cont~lner4. ~b~e contAlner~ are more g~ per~nble th~n the "P~r~t g~ner~tlon~ of blood ator~ge contalners that are ma~e of polyv~nyl chlorlde ~P~C).

~he u~e o~ these opecial contalner~ a110WB cArbon dloxlde ~C02) ga~ to e~cAp0 durlng ~torAge of th~ blood component. The e~cap3 of Co2 g~ roduces the fo~atlon of carbonic a~d ln the prewrvatlon of stoe~ge ~olutlon. ~he roductlon of carbonlc acld ln 601utlon extend6 the per~od of tlme durlng ~h~ch A phy~lologlcally ~c~opt~ble pH c~n bo m~lntalned ln th~ stor~ge ~olutlon.

U.B.

Pat-nt Numb~ 4~447~415 ~o Rock et al.

d1BC10~e8 A llqu~d ~torage m-dlum for pl~telet8 tbat 1B pla~a-frce. The m~dlum o~ thls lnv-ntlon U~B one or more addltlv~ ln oon~unctlon wlth a rallne and antlcoagulant, d~xt~o~e-contalnlng ~olutlon th~t 1d~lrably a form o~ CPD-~yrode' ~olutlon. ~h~ Addltl~eo dlsclo~od a- b-lng ~u~table for u~ wlth thl6 lnv-ntlon ln~ludo ~1) e~or~lblo lnhlblto~ that are 13264~0 organlc compound¢ such ~ lndomethacin, quinPcrlne, or vltamln E and ~2) ~ub~t~nce~ to ral4e oyclic Adenoslne monopho~ph~e levols ~uch A6 prostagland~n6 El, ~2~ oe . ~any of tho6e addltlves fall ~o m~et ~afety and regul~tory requlrements required for ~u~t~nces ~or ln~u610n lnto human~ and ~re, therefore, only ~ult~bl~ for expe~imental us~ or only f or ln ~t~o uao~ Other addit~v~ di~clo~od ~ ~ul~ble for U6s w~th thls invention include ~1) nutrlent~ such ~8 ~ructo~e ~nd ~ther ~ugAr6, adenlne, or ~¢etyl COA ~nd ~2) buffer6 6uoh ~6 phosphate and certaln ~mlno ~lds. The org~nlc compounds o~ Addltlves ldentlflod a~ nutrient~ do not oll~in~te tho requlrement for tho p~e~ence of dext~o6e ~ n tho modlum, The combloatlon o~ additive and gluco~ oarrlod ov~r ln the pla-ma ~atl6P~06 th- nutrlent requlrom~nt ~or tho plat-lets for porlods o ~tor~qo tlme cxtond~ng up to 5 days ~nd tho Addltlves ldonti~led ~ bu~er~ cannot malnt~ln ~ balanced pN durlng oxtended plAtolet ~torage p~r~od6 boyond thls tl~. 81nce the ~lu004~ lovolr are clearly ln~do~u~te ~or r-d coll ~torA~ ovor longer tlm perlod# ~nd these buff~r~ c~nnot ~doquAtoly bu~er the ~ount o~ lactlc oold produced by vlable, ~opon~e~ ~od c~ o~orod ~t temporAtu~B o~ 4 1 2C Ccll deAth due to pH ~All lo llk~ly, ~ a re~ult thl~ solutlon, though potontlally 132~ ~cceptable or pl~telcts, c~nnot be used to ~ore rod ~ells. Thi6 ~i~elosurq ~, the~fore, not dlre~ted to storage medium for red blood cell~.

~ he lndustry i~ lack~ng ~ 4100d coll ~torage odditive med~um that 1~ free o~ pla~a and orgAnlc compound~, which c~n be used to ~tore red blood cell4 and platolet~ for extended p~iod# of time an~ wh~oh 1 6~e fo~ in vivo hum~n u~e.

~_ .. _ .

BRI~ DESC~IPTION OF T~E FIGuR~S Flgure 1 gr~phically compAreg the peroent los~ of ~P in r~d blood cell~ After ~g days o~ storage ln va~lou~ embodlment~ of t~o rod blood cell ~torage ~dium ~nd ~ control ~tor~go modlum.

Figuto 2 gr~phlcally comp~re~ the percent of hemoly~ls o~ rod ~lOOd cell~ ~fter q9 days o~ ~tor~ge ~n v~riou~ embodim-nt- of the rod blood cell ~torAgo medium And ~ control ~torAg~ medlum.

Flguro 3 graphlcol~y compare~ the p~ o~ ~toroa ~ed blood oell ln 801utlon ~ft0~ 49 day~ o~ stor~go ~n v~e~ou~ o~bodimont~ o~ tho r-d blood ooll ~torAg~ modium.

Figuro 4 gr~phlcally compAr~ ~ho porcent 10~8 0~ 2~3 DPG ln rod blood cellr aft-r 4~ d~y~ o~ rtora~- in -13 13264~0 v~riou~ e~bodim~nt~ of th~ red blood cell ~torage med~um And u control 6tor~ge mediu~.

SUMMA~Y 0~ TI~E INV~NTION Thl~ lnvention 1~ a sterlle, pla6~a-free Gtor~ge med~um for blood component~ ~nclud~ng red blood cells and/or platelets processed separately or together. The mo~t de~r~ble embodl~ent of thl~ lnvention i8 a ~terll~, plasm~-free red blood c¢ll 6tor~ge med~um. Th~ red blood cell s~or~ge medium in~lude~ a phy~lologlc~lly comp~tlbl~, ~queou8 ele¢trolyte s~lutlon. ~n one liter o th~s electrolyte solution there 16 bet~e~n about 3.0 ~r~m~ ~nd ubout 25.0 gr~ms of dextroRe, betw~n about 3,0 gr~m~ an~ about 6.0 grnm~ of ~odlum citr~t~, and between A~out 2.0 gr~m~ ~nd about 4.2 g~am~ of ~odlu~ blc~rbon~te. The red blood ccll 6torage medlum l~ 160tonlc wlth the blood to be stored and ha~ a pN ~n a r~nge of between about 6.~ and ~out 7.4. ~he rod ~lood cell 6tor~ge medium 1B capable o~ ~to~ng and pre~ervlng red blood cells for ~t 1ea~t 4g dAys at ~ tempe~ature of ~t least ~bout 4c. ~he embodlments of the ~nvention that are most de~lr~ble ~nclude about 2? mg/dl o~ ~denlne.

~ he red blood cell ~tor~ge medlum can have electrolytes includlng~ ~n l llter of the medi~m, -14 132~0 bçtwoen about 6.6 grams and about 7.6 gram~ o~ 60dlum chlorlde, b~tw~s~ about 0.2 gr~ ~nd abQut G.4 gr~ o~ pota~ium chlorld~, between about 0.1 gram ~nd a~out 0.4 gram of salelum ehlorlde, botween about 0.2 gram an~ about 0.4 gr~m of magneslum sulph~te, ~nd between about 0.1 gra~ ahd about a . 6 gr~m o~ monoba~lc godlum pho~phate.

~ he lnventlon ol~o lnclud~ ~ p~oce~ for preser~lng red blood coll~ ln a sterilo, plasma-~ree ~tora~ med~um.

DETAI~D DE5CP~P~ ~C3~3=~Y Thl~ invent50n 1B ~ sterllc, pla~a-~ree stocage med~um ~or blood components lncludlng red blood cell~ and platolot~. The most de~lrable cm~odlment o~ th~E lnventlon ~ 6 a 6t~r~1e, plAsmA-~ree red blood cell ~torage medlum. Tho red blood cell 6torage med~um lnclude~ A phy~lologl~lly compatlble, aguoou~ ~loctrolyto w lutlon. ~n one llter of thls eloctrolyte ~olutlon ~hero 1~ botween about ~.0 grams and about 25.0 gr~ms of glucos~ or more do~lrably, dextrose, between about 3.0 gr~ms ~nd ~bout 6.0 gr~m4 o~ ~odlum oltrAt and b-twoon ~bou~ 2.0 gram~ and about ~.2 ~ram~ o~ ~odlum blcarbona~o. Th- r-d blood coll ~torago ~odlum 1~ lsotonlc wlth hum~n or anothee blood to bo ~tored and -15_ \~/ ~J - 1326~40 hAs ~ p~ ln a r~ng~ o~ betwsen about 6 ~ ~nd ~bout 7 4, ~xcept for the nutrlent ~ugar th~t 1~ deslr~bly dextrose, citrlc ~cld or cltrlc ~oid deriv~tlve~, sodlum blcarbon~te And ln cert~ln embodl~ents~ ~denlno~ the red blood cell ~orage m~dium of thiB inventlon 1~ f~ee o~ oeg~ni~ compound addi~lves ~he term "vl~ble" red blood cells ~8 usad here~n mean8 that ~ub~tantlAl concontratlons of the l~olAted platelet~ ~uspended in the red blood coll storage mo~lum retaln thelr normal and ~nherent phy~lologlcal, functlonAl, nnd structur~l properties such that the stored r~d blood c-lls ar~ ~ultAble for lnfuslon and functlon after lnfu610n ln a reclplent Th- phy~iologlcally comp~t~41~, ~queou~ loctrolyte Eolutlon Or th~s lnventlon can ~e ~ar~d wlth only m~rglnal ef~ect on the ~tor~go c~pab~llty of ~h- red blood c-ll ~torage ~edium ~he most deslrablembodlments of the red blood cell stor~ge medlum conta~n th~ mo~t ~lgni~l~ant olectrolytcs ~ound ln blood pl~m~ ~h~ electrolytQs are contalnod in th~ red blood cell storago m-dlum ln the s~mo ~pproxlm~te conc-ntr~t~ons ~B ~ound ln nor~ lood pl~oma ~h~ ~ost d-~lr~blo electrolyt-~ lnolud- odlum chlorlde, pota~slum chlorld~, c~lclum chlorld-, m~gno4~um ~ulphat-, and monob~Blo ~odlum phorphate / - 'J `J 1326~0 ~ lectrolyte~, such ~B tho~o ldenti~le~ ~bove ~nd othor~, ~re com~only ~v~ bl~ in ~queoue ~olutions ~or ~n~ectlon or lnfuslon into a ~eclpient. ~n prep~rlng th~ red blood oell 6to~gc ~odlum the concentr~tion of the~e ele~troly~e~ ean be ~ltered by known technlque~ to obt~ln ~n isotonlc ~olutlon. A deslrable ombodiment of the r~d blood c~ tor~ge mod1um h~ eloctrolytes whlch ln~lude, ln 1 lit~r of the medlum, between about 6.~ gr~ nd About 7.6 grams of sodlum chlori~e, botwe~n ~bout 0.~ gr~m ~nd ~bout 0.4 gram o~ pot~6~1u~ chlor~de, botwo~n About O .1 gr~m ~nd a~out 0.4 gram of calclum chlorlde, bet~een about 0.2 gr~m and about 0.4 grA~ of m~gno~lum ~ulphat~, ~nd between ahout 0.1 qr~m And about 0.6 gr~m o~ monoba~lc ~odium pho~phate.

A do~lr~bl~ stor~g- ~olutlon for r~a blood cell~ 1 ono th~t cont~inr eloctrolyt~s that mo~t closely ro~o~blo blood pla~m~. An lonlc ~nv~ronment ~lmul~tlng proto~n-~roo p~aBma producc le~8 msmbr~ne p~turbAnce and he~oly~i~ ln ~tored r-d blood ~ , Th~ more ~t~ble mombr~no improv-r th~ norm~l m~tabollam o~ the r~d blood c~ uch th~t A~ ~nd 2,3 DP~ ~r~ m~int~ln~d and lactat~ output lr r-duced. Normal mctabollrm le~en~ tho n~od ~n ~torAg- olutionr ~or ~pcc~a~ r~d blood c~ll add~t1ve~ ~uch ~8 ~d~nlne, mann~tol, ~nd h~gh ~luao~e.

-17 ~, 'J ~326~0 G1UCOBe ln the form of d~xt~oge ~ 4 th~ natur~l nutr~ent for red blood cell~ ~extro~e 61gn~flc~ntly contribute~ to the ~billty of th~ red blood cell storag~ m~iutn to pre~rve an~ m~lntaln v~ able red blood cell~ for extonded ~torage period~ beyond a 35 d~y perlod The r~d blood cell ~tor~ge medium of this lnventlon u6e# dextro~e n~ tb~ only ~igniflc~nt nutrlent foe the ~to~ea red blood cell~ An ~nslgn1 flcAnt pre60nce of another nutrlent or the u~e o~ racRmlc gluco~e doe6 not ~pprecl~bly ~lter the effectlveness of the red c~ll ~tor~g~ med~um of thl~ lnventlon The pre~ence of ~noth~r nutrlent 1~ undeslrablQ because other nutrlent~ ar- not AEI effectlv~ ~8 dextrose ln the lonq t~r~4 pre~ervatlon of red blood cells The conc-ntrAtlon of daxtro~e ln the r-d oell fitorag~ med~um o~ thl~ ~nv~ntlon 16 th~n the daxtrose cono~ntr~tlon~ typ~cally u~od ~n 601utlons for uBe wlth whol~ blood or blood co~ponQnts. ~he hlgher doxtro60 ~onc-ntratlon must be ~f~lolent to p~ovl~o nutrlentr ~or the ~tored r-d blood c~llq throughout thele torAg~crlod Pextror~ $~ d~lrAbly pre~-n~c ln th~ rod o~ll ~tor~go mcdium ln a ~on~-ntratlon of ot ~eP6t ~bout 3.0 gram~ per llt-r and mo~t del~lrAbly at about 7.2 qramr p-r llt-r Typl~all~, a conc~ntr~tlon of d~xtro~e of b-tw--n about 4.0 to about 25.~ gr~m~ p-r llt-r 1~ --18_ - ~3264~ ~uEflolont to provlde r~d hlood cellR, ~tored accordlng to thls lnvent~on, ~lth ~u~flcient nutrlents ~or At len~t About 4~ d~ys. ~h~ oonoentr~tlon o~ dextro~e ~n embodlment~ o~ thl~ invontlon u~eul for the stor~go of platole~ at lenst ~bout 3.0 gram3 pQr llt~r and de61rably ~etwotn ~bout 3.0 grAm~ and 7.S gram~ per l~t~r.

The buffer ~y~tem of the red oell 4tor~go m-dlum o~ thi~ lnvention 1~ cr~tic~l to tho succe66ful 6tora~o of red hlood oell6 for at l~a~t about 4g day~. ~he use of sub~tantlal concentration of sodlum bi¢~rbonate to the red c~ torage medlum n-utrall~e~ much of tho laotic ~o~d fGr~d during red blood cell ~torage. Tho ooncen~rat~on o~ ~odium blcarbonato mu8t bB ~uffloient to malntaln n rod ~oll storage m-dlum pH ~or the ~tored red blood oell4 at ~bov~ about 6.5 throughout the torm of the ~torage perlod. A r-d oell storAge medlum pH v~lue bolow AboUt 6.5 cAn b- damaglng to t~e tored red blood ooll~ ~r ro cted ~y ln ~lt~o p~rAmot-r~. shO~0 ln vltro par~motor~ Include ~1) r-d blood coll m-mbrano ~k-l-ton ~ltoratlon~ r~sultlng ln dearo~sod deform~ ty and lncr-asod fr~glllty or hemolysl4, (2) th~ d~plo~on of ATP, and ~3) lo~ o~ 2,3 DPG 10~01B r-~ultlng ln ~ltor-d oxyg~n-trans~r ~hAr~ctorl~t~cr.

~o~oarch ha~ domon~trated that tho pH l~v-l of tho -19 ~ J 1326~0 ~to~d red hlQod cell6 cAn be m~lntalnqd and th~ ~,3 ~PG level ~abilized by the Addlt~on of a hi~h cono~ntr~tlon mM) of ~od~u~ ~icar~on~te to the rod cell ~u~pendlng medium Ths u~e o~ ~uch A high, nonphy~iologlc sodium ~lchrbonote conc~ntr~tlon c~n requlr~ the presence of ~denlne and mannltol ln the 601ut~on to prevant metabollc and membrane dlsturb~nco~ Th~ bu~fer ~y~tem o~ th- red cell ~torAge medlum o~ thl~ lnv~ntion us~ sodlum bioarbon~te A~ the pr~nclp ~lk~llne agent 80dlum blcas~onhte i8 used ln concentr~tlon~ sufflolent to mAintAln the do~lred pN vAlue of the r~d ocll 6tor~qc ~edium throughout red blood cell storAge wlthout preclplt~tion of tho buff-rlng ~qont. Do~lrAbly~ ~odlum b~CA~bonAt- ~B pre6ent ln 1 llter o~ red c-ll 8tor~90 ~edlum at a conconts~tlon of betwecn about 2,0 grAms to about 4 2 gr~m~ The bu~or ~yrt-m o~ th~ red c~ torAge medlum of thl~ lnvontlon de~lrably lncludo~ ~onob~-lc ~odlum pho~ph~te M~nor concontratlon~ o~ other ~alt~ c~n bo sultab~o ~or lnclu~lon wlth tho bu~terlng ~y-tom o~ thl~ lnventlon, ~ ho ~ntlcoagul~nt ubed ln the red oell 6~0~a9e modlum o~ thls lnvcntlon lnc~d~s ~odlum oltr~to. ~n th~ mo~t prO~eFrCd ombodlment~ of the lnventlon cltrlc Aold 16 lncluded. ~h- ~ntlco~gu~Ant~ of thl~ lnv~ntlon -~0 132~0 ~u~t ~ pro~nt ln oonconteatlon~ ~u~flGlont to provent ~ubst~nt~l co~ulatlon o~ rod b~oo~ c~118 durlnq oxtond~d storage per1ods. ~o~lr~ly, ~o~lum al~rate 1 p~ n~ In l lltor of rod blood coll ~to~Ag~ ~dlu~ ~t concentratlon o~ ~otwo~n ~bout 3,0 gr~e And about 6.0 gr~m~ ~nd oltrla ~ld 16 pro8~nt ln I llt~r of red blood c~ll storago medlu~ At ~ ¢oncontr~tlon o~ botween ~bout 0.~ gra~ and ~bout 0.6 grA~.

Mlnor ~oncCntrAtion8 o~ other ~ntloo~gul~nt~ cAn be sult~blo ~oe incluslon ln the rod b~ood coll ~toraq0 ~dl~ of thl~ inventl~n~ ~ he rod blood ooll ~torAg~ mQdlu~ o~ thl~ lnvontlon 18 d~lrably ~ompo~od of th~ ch~mlc~l lnq~cdlonts ~18t-d In ~able S. Tho most do~irAble ombodl~nt~ of tbl~ ln~ntlon ~on~l~t 0~60n~1Ally of tho~o compounds to tho oxclu~lon of ~ sl~nl~lo~nt concentr~tlon of any othoe com~ound~ The o~clu~lon o~ othor compound~ ln th- r-d bloo~ cell ~tor4g- ~o~lu~ o~ thl~ lnvontlon 1~ dorlr~blo ln ord~r to pr-~ont r~n~ltl~tlon ln th~ r~cl~lont ~nd to ~xlmlro the ~torAg~'perlod ~or rod blood c-llo -21 1326~ TARI.E 5 Concent~atlon Preferrod Ran~e _ _ concentratlon Sod~u~ Chlorlde C.4 - 7.6 g~l 6.450 g/ Pot~8~ium Chlorld~ 0.2 - 0.4 9/1 0.375 g/l C~lclum Chlot~de 0.1 - 0.4 g~l 0.248 g~l ~agne#lum ~ulfate 0.2 - 0.4 g/l o.aoo g~ ~odlum Pbo~ph~te ~monob~c) 0.1 - 0.6 g/l 0.355 g~l ~extro~o 3.0 -~5.0 g/l 7.2 g~l c~tri~ ACld 0.~ - 0.6 9/l 0.510 g~l Trl~8Odlum Ci~r~te 3.0 - 6.0 g/l 4.471 g/l 8Odlu~ ~l¢~rbonate 2.0 - 4.2 g/l 3.000 g/l Adenino 0.26- 0.2ag/1 0.270 9/1 ~h~ ~n~vldu~l lonlc ch~r~cter of the rolutlon ln mEq~ a# ~ollow~ ln ~abl- 6s -22 1326~0 TW LE 6 ~nge ot ~onlo Pr~f~rred ~onlc Conc~ntrQtlon Conc~ntratlon N~l ~S~.B0 - 236.10 207~60 . 2.6~ - 5.36 . 5.03 C~+~ 1.36 ~ 5.45 3.38 Mg~ 1.70 - ~.40 1.70 NP0~ 2 1.38 - 5.50 2.58 Cl- 1~0.00 - 138.00 117.38 8O~-2 l.?o - 3.40 1.~0 - 23.80 - 50.00 35.71 ~ bo p~ of the ~d blood coll ~toroqo ~edlu~ 1~ ~alnt~lnod ln ~h- rongo o~ 4-t~oon obout 6.8 ~nd ~bout 7.~. Tho pH o~ tho ~to~og- ~odlu~ ~n v~ry du~ to th~ A~ount o~ CO2 pro~ont ln tho cont~lno~ ~ hl~h concontratlon o C02 lowor th- pH o~ tho ~tor~o a-d~u~.

~ho hlgh pN o~ tho ~tor~-.modlu~ c~n roqulr6 ~op~to ~t~rllle~t~on o~ tho d~xtro4- rom tho ~od~u~ blc~rbon~to-cont~lnlng oto~a~o ~odlu~ ~o~u-- d~xteo~le not eo~dlly ~t~rlll~od Pbov~ p8 6.4..

Tho b~lo ~olu~lonr ~nd lngrodl~nt4 ~ult~bl~ ~o~ u40 ln ~Pklng th- r-d q~ tor~go ~od~u~ o~ thl~ lnvontlon o~n b~ obt~lnod ~ro~ nu~orou~ oo~morclal -23 ~326q~o BourceS a~ ster~l~, non~pyrog~nic, lnicct~ble ~olutlon~.

Suppller~ for th~ lngredlents c~n bo identlfio~ feom common publio~tlons ~uch a~ the "Physlcl~n'~ ~o~ Reference~ ~nd the "Red Book" e~ch publi~hed by the Medical Econom~ce Comp~ny ~nco~por~ted, o~ad~ll, New Je~ey. ~he ~ollow~ng ex~mple~ of ~omme~ lngeodients are provlded ~8 ~ ~mpllng of ~ccept~ble commercial product# av~llable ~or u~e ln thi~ lnventlon.

~lngQr~s In~ection, U~P, contAln~ 8.6 ~ram~ of sodlum chlorid~, 0.3 gr~m of potasslum Ghlorld~, ~nd 0.33 gr~m of c~lcium chlorlde per llter ln sterlle, non~pyrogenlc w~te~ for in~octlon. Sterile w~ter for in~ectlon 1~ a non-pyrogenic w~ter for lntravenous infuslon. Magne~lum Sul~Ato In~octlon~ U~P, 1~ a 50 percent solutlon o~ magneslum ~ulph~t~ ln sterll-, non-pyroqenlc wat-r ~or ln~c~tlon. sodlum bloarbonate ln~ectlon, USP, 1~ an a .

percent aoluelon o~ sodlum blc~rbon~te in ~terlle, non-pyrogen~C water for ln~ection. Dextro~e ln~octlon ~olutlon, USP, ~6 S~ percent ~olutlon of dextrose ln ~terlle, non-py~ogon~ water Soe ln~ectlon. Yota~lum Chlorldo ~n~oct~on, U8P, 1~ a 22 p-rc~nt solutlon oS po~aa~lum ohlorlde ln t-rlle, non-pyrogonlc wator ~or lnioctlon. An~looagulant citrat--phosphat~-d-xtro~olution haB 3,0 grama o~ cltrlo acld, 2~.3 gram~ o~rodium citrato, Z.2Z g~a o~ aodlum blphoaphato, 25.5 -24 ~._J V 132~4~0 gr~m~ of dextro~e ln one llter ~terlle, non-pyrog~nlc wator and lB u~ed in the proportlon of 70 m~lllllter~ of CP~ to 500 mlllil1tor~ of wholo blood Ad~nln~ 18 availa~le, from b~th ~yn~hetlc ~nd nat~ral ~ouro~, aB a powder Tho above list~d ingredient~ ~re combin~d ln the proferr~d embodiment of tha lnventlon in the followlng amount~ ~he rGd oell ~torage medlum solutlon contaln~ 750 mlll~liter~ o~ Rlnqer's ~olutlon, 170 mllllllters o~ ~PD, 40 mllllllter~ of sodium bic~rbon~te solu~ion, 5 4 mlllllleer~ of dextros- in~ectlon ~olution, 0 7 millll~tor of po~assium chlorlde, 0 4 mllliliter of magnc~um ~ulphate solutlon, 33 5 mllllllt~rs of storl~ B wator for ln~sotlon, ~nt 270 milliqram~ of ~denlns All ln~redlent~ ar- combln-d undor sterll~, aseptlc contltlons Prlor to lnnocul~tlon of rod blood C0118 wlth tho eed oell torag- modium, the mlxtur- of thlngr~dl~nt~ ls flleor ~terlll~d us~ng a C Z mlcron fllt-r unlt de~gnod for the v~cuum flltratlon o~ ti#~Uo ~ulturo modl~ ~ ho pr-f-rr~d eo~porltlon o~ th- rod c~ tora~mo~lum o~ th~ pr~en~ lnv~ntlon and cP~-pla-ma aro~mparod ln ~abl~ 7.

--25- 326~0 ~ .

R~d ~ll ~ CPD-P~ m~ tq~l ~odl~m 2n7.60 173.0-180.0 POtABl~lUI~I 5 . 0i~ 1 . 9--3 . 8 ~lc~ a `3 3~ 3 ~ o Magn~Blu~ O ~ -9 Pho~phat- 2.S8 3.g Blcarbon~t~ 40.00 20.9 Cltrat~ ~S.00 22.~ Bulphat~ 1.7~ 0.4-1.1 ~hlorld~ ~17.3~ 75.0-~0.0 Prot~ln~ 0.0 12.2 Orqanle Acld~ . 4-4 Cltrl~ ~old2.~0 ~ol/l 3.9 ~nol~i D~xtro~ ~0.00 m~ol/l 25.0 ~ol~l ~d~nl~o 2.00 ~oljl b.~ ~ol~l ~ hl~ concont~tlon a~u~ th~t th- cltrat~, olt~lc ~old, ~nd pho~ph~t~ do.not ~nto~ rod aoll- and 1~ ba~d on ~ 70 ~llllllto~ CPD antlooagul~nt ~olutlon ln 500 m~lllllto~ o~ wholo blood havlng ~ h-~toorlt o~ ~2,5 and ~-rum ch~loal ~alu-~ wlthln th- no~ lt~ A~ ll~t-d ln H~ri-r'~ R-vlow o~ ~looh~ tr~.

-2C . .

~326~ ~ ft~r the pr~paratlon o the red blood e~ torag~ medlum the proc~ OF pro~ ln~ And storln~ r~d blood c~ r~qulr~a th~ 6~p~ratlon o~ r~d blood ~ feom th~ oth~r compo~nt~ o~ blood. ~h~ pl~s~ oxpre~s~ off~ and ~ollected ln ~ ~telllt~ bag. Tho red o~ll stor~g~ 1u~ can ~hcn b- t~an~f~rr~d lnto the cont~lnor holdln~ th~ r~d blood ec~l~. ThlB trAn~f~r ~n bo by al~hsr eonnectlv~ tublng llnklng tho ~at-lllts b~ oontalnlng the r~d eell ~torag~ ~edl~ or oo~mer¢l~lly av~llablo, ~t~rlle oonnoetlon dovleo~ th~t e~n ~r~n~or th~ ~odlu~ ~ro~ a oontalnor not orl~l~ally attaoh~d to the collectlon bag 8et~ Aftor rorurpon~lon of th~ 6ep~r~t-d r-d blood coll~ ln the rod ~loo~ ¢oll ~tor~go ~odlu~, th~ r~d blood e~ ro otorod ~t ~C.

Convontlon~l P~-732 r-d blood ~oll ~ont~lnoe~ h~o a hlgh p-rm-~b~llty to Co2 8torago of tho ~od blood oel~ ln g~ porm~blo oont~lnor~ ~uoh a~ co~rcl~lly ~ bl~, thln pol~vlnyl chlorldo (PVC) baqr Allo~ the o~o~p- Or C02 ~nd, th-roby, lo~on~ eho n~od fo~ ~od~ ~lo~rbon~t~ ln tho r-d o~ tor~ o~lu~ ~hl~ 1boc~u~o th- dl~fu~lon ot C02 ~ro~ th- contAln-r d~or-Astho ~m~unt of carbonlc acid r-t~ln-d ln tho oont~ln~d ~olutlon, Sn pl~tol-t ~tor~go tudlo~ u~ln~ a plAt-l~t cont~ln-r, 40~M o~ blc~bo~Ato ~r outtlcl~nt to n~utr~ o 30~ o~ l~otlo aold to,Aohl~v- a ~lnal ~H o~ ` r~ 13264~ 0 more thAn 6.8. A pH of 6.8 can be expe¢'ced to ~ccumulate ~ ~bout 42 d~y~ o~ ~tor~ge o~ rod blood cell~ at 4C.

The red blood cell s~orago medlum o~ thl~ lnvent~on a110WB for 6tor~ge ln ~hln polyvlnyl 6torAge cont~lner6 pl~6tlcl~ed wi~ d~ethylh~xylphth~l~te ~DEHP) ~uch ~6 ~he C02 permeable cont~ln~ u~d for the ~to~Age of pl~tel~t~. Tho6e contAin-rs le~sen the need for sodlum blcarbonate becau~e ~ore Co2 g~ leave~ th~ con~inor and le~s carbonlc ~cld 16 di~OlVCd ln the ~olution. ~he pre~ence of D~P a# ~ pla6t~cl~er ln ~ cont~iner c~n enb~nce t~e qu~llty o~ ~tored red blood cell~.

In ordor to demon~t~te tho ~u~t~bility o~ the ~tor~go modlum o~ tho pro~ont inv~ntion ~or pre--rving blood co~pononts, ln vlt~o ~nd ln vlvo ~tudles w-re conducted to comp~re the qu~lity o~ platelets concentrate~ torod la th- ~torAg- medlum wlthout tho peerence o~ ~denlne or ~nother membr~n- st~blllslng c3mpound. ~x~plo~ 1 through 7 ~th tholr r-l~t-d Comp~tl~o ~x~mplo4 A th~Qugh I prorent the r-~ult- o~ tho~o t~t~. ~n tho~ ~x~mplo~ ~nd Compar~tlv- Sxampl~th~ storage m~dlum W~B ox~mln-d ~or lt~ u~fuln~r~ ln tor~ng platel-t~ and 1- d-~lgnat~d by th~ t~rm~ NPl~t~lot 8tor~g~ M-dlum" o~ "P~8.M.

-2a- ' r~~.

~ v 1326~ ~ho ollowing zxamplo~ ropr~ent tho inv-ntlon.

The comp~r~tlve ex~mpl~ ~o not repr~s~nt the lnventlon, 15XAMPliES 1 TNROU~H 5 AND ~ h~ proceduro u~d in the~e ~xA~ple~ ~n~ ~ompar~tl~e examplo~ to eeparate plAt~let~ and to ~ake th~ Btor~ge mod~um WAB ~8 dQ~ori~od ~bove and $n th~ parent app~i~atlon, now U.~.

Patent Numbec 4,695,460, for t~e p~o~rred smbodlmont o~ tha platnlot invent~on.

Tho daSo pr~sent~d for tho platel~t stor~g~ medlum 16 desl~n~ted b~ th~ ~ymbol "P.~.M.~ and reproBentB Ex~mplos 1 through 5 td~ na~od by th~ 8ymb01 "Ex.") ~or the lnvont~on. The dat~ pro~ented for tho ~torag~ o~ pl~tolot~ ln CPD-pl~m~ ~ de~lgnat-d by th~ symbol "CPD-P~ nd rQpr~ents Comp~ratlve ~xampl~td~lgn~tod by th~ ~ymbol C.Ex. n ~ A through B.

For the~o ox~pl~ and c~mp~ratlve ox~mpl~plat-lo~ ~oro B~parAt-d~ 4torod ln thelr ~o~p-ctlv~ ~edla, and t~tod on day~ , 10, ~nd 14. Th- te~t~ condu~Sed on th--- day~ ~or plat-let count d-tormlnod a porcont o th- platolot count o~ tho flr~t day, th~ per~ont lncroas- ~n optlc~l donolty ~0.~) o~ tho oxt-nt of plat~lot sh~po chang- wlth ADP, tho porcont of hypotonlc hock ro-pon-~, tho conc-ntr~tlon of adono~ln~ -29 ~ ~J ~3~4~ trlphosphat~ or "~TP" in th~ plAtelet~, And the DF~ount of ly818 1~1~ evldenced by l~tAt~ dehydrogen~40 or "r.DN" r~l~a~ed ~y th~ pl~lt~l~tB.

Th~ results of the8e to4 ~eo re~pectlvely pr~ented ln TAbles 8 through 12.

ABLE 8 Pl~tole~ C~ * DAy 1 DI~V 5 I~ y 10 EX. 1P.S.M. 100 94~3 91~2 B2~4 C .~X. ~ CPD_P1. 100 94~3 84 16 77~3 B~ g ~Xtent Of Sh~P~ ChAnge~ ~ InCre~Be 1n ~.D,~ ~ DAY 5 DaY 10 P~v 14 ISX. 2 P.8.M. 16~1 14~1 10~1 7~1 C.Ex. ~ CPD-pl. 16~ 1 5~1 4~1 A~E 10 ~ypotonlc 8hoak ~sPon~, %~ D~y 1 ~ DaY 10 D~ EX. 3 P.B.M. 7516 65-l4 63~5 40+3 C.Ex. C CPD-pl. 82~3 7B~5 49~2 12i5 ~A~LE 11 Aq~P, nmolo~ oll Vltll .

D~Y 1 D~Y S 32~L~ ~ ~X. 4 ~.8.M, 8.St~0.4 7.7~0.3 5.4~0.4 3.1~0,4 C.EX. D CPD_P1. 7.S~O.7 6.0~0~4 3-S~O-3 0.8~0.2 _~o_ ~J ~J ~3264~0 TABLB 1 2 L~H, Un,1t~ R~1e~edb aY_ 1 DaY 5 D~V l O PAY 1~ ~X. 5 P.8.M.138~16 177~22 305133 430~43 C.EX. 15 ~::PD--pl. 120+8 215+15 430+23 585:F~5 *~e~uIt~- represent the meAn + standard devl~tlon Th~ r~sylts of th~e comparatlv~ ~tudle4 domon~trate that the plat~lets 6tored ~n the pl~telet ~torago medlum ~howed better malntenanoe o~ ~orphologlG And phy~lologlc lnt~grlty a~ lndlcated by the ~ollowlng The platolets ~uspend-d ln th- plAtalet ~tora~o med~um of thl~ lnvcntlon d-mon~trated a bete~r pro~orvatlon ovldonood by the dl~fer~nc-- ln pl~t~lot count ov~r th~ p~rlod of te~tlng A docroa~e ln pla~elot count r~fl~ct~ plat~l-t olumplng and/or ly~l4, ~ho pl~tolotæ ~u~p~nd-d ln th- platcl-t ~torag- m~dlum of thl4 ~nvcntlon demon~trAeed a bott~r malnt-nanc~ o~ tho ablllty o~ th- plat-l-t~ to und~rgo shap~ chan~o or to b-como actlvat-d by u~ln~ phy~lologic actlvator~ Thplat~l-t~ ~u~p-nd-d ln th- pl~lot ~torage m~dlum o~ this lnv~ntlon d-mon~trat~d a bottor pr~so~vat~on than tho pl~t-lct~ ~u~p-nded ln CPD-pl~a by th~lr ablllty to r~cov-r from hypotonlc ~tr-~s Tho platolot~ u-p-nd~d ln th- plat-l~t tora~o modlum o~ thl~ lnv~ntlon dc~onrtr~tcd a b~tt-r m~lntonanco o~ tho A~P -31 r~ ~, w ~ 132~4~0 l~els whlch roSl~ct~ tho onorgy 6t~tU~ 0~ th~ pl~telot c~ ho pl~tol~t~ ~uspondo~ ln th6 pl~telot ~tor~g~ m~d~um of thl~ lnvontion d~monstrot~d ~ bott~r ~nten~nco of m~mbrAna lntogrl~y ~4 ln~lc~tod by le~ lo~ of lntr~c~llul~r ~DH durlng 4toro~0 .

Thoso ox~plo6 ~nd comp5~tlvo ~x~pleE d~mon~trat~ ~h~t 8torag~ of plat~l~t con~entrat~ ln th~ pl~tele~ ~tora~ dlu~ for ot l~a~t $0 to ~ dAys At non~r~ozlng to~poraturoll or o tomperotueo o~ ~t l-~t about 2ac m~lntAln ln ~ltro qu~llty that 16 ~f~c~lve of ln vlvo vloblllty, elnll~r to thot obtaln~d wlth storAge o~ plot~lot~ ln CPD-pl~ or 5 to ~0 dAy~ .

~XAMPLE 6 COMPWtA~IV~ MPt.t8 1~ OU~N H Thl4 ox~plo ond co~p~r~ o ~x~pl~- u~- th~ ~mo proooduro ~ do~orlbod ~o~ ~x~pl~ 1 through S and Co~p~ratlv- ~xa~pl~ A through ~. Shl~ ~xa~pl- ~n~ oo~p~ratlvo x~mplo~ d~on~tr~to thc ott-ct ot u41ng vaelou~ a~ount~ o~ ~odlum olt~t~, 60~1u~ chlo~ld-, m~gno~lum ~ulph~to, ~o~lu~ dlpho~ph~t~, ~odlu~ blo~bona~-, PC02 ton~lon-, d~xtro~, an~ pl~ ln dlff-r-nt pl~t-l-t ~tor~go modl~. shcs~ ~odl~la~tlon4 ~r~ on~trat~d by oo~p~rlnq ~ ct~ ot dltt-r~nt pl~tol~t 4torAgo ~dla ovcr a lO d~y ~tora~ po~lod on 132~0 ~1) pl~telot co~nt, (2) the p~rcent o~ hypotonlc shock re~pon~o, ~3) th~ ~tructural ~nt~gel~y o~ the pl~tolo~# ch~rao~erlzed by chang~ ln ~l~e dlstrlbutlon, Appoar~noo o~ pl~t~le~ clump6, balloon formo, fr~gm~nt~ ~6 ~dged by mlcro~copy, and LD~ releasa, t~ pl~t~l~t functlon a8 ¢h~r~oterl~ed by th~ extent of shap2 change wlth ADP, ~nd (S) platel~t ~nergy m~taboll~m or the rat~ of ox~g~n uptake, lactat~ productlon, glucos~ eon~umption, and ATP level~. The d~t~ demon~tratlng t~o~e ohar~ct~rl~t~c~ ~re pre~ented re6p~ctl~ely ln ~ble~ 13 through ~7, Exampl~ 6 pr~nt~ data ~or the ~Ive char~cterlo~lc~ for th~ plat-l-t ~torag- medlum o~ thl8 ~nvcnt~on ~nd 18 do8~gn~ted br th~ 6ymbol ~P~S~N~ ~ ho compArAt~v- ~x~mpl-s pr--ant d~ta or tho flvo ch~ra~t~r~ tlc~ for th~ platcl-t storage m~dl~ d-~lgnatod by th- ymbols "~M", ~BSM ~ gluco~-~, and "DMSM". Tho ~y~bol "BSN" ropro~nt6 ~ ~torago ~odlum h~vlng th- ram~ characteri~tlc~ of th~ pre~-rr-d plat~l~t ~tora~o m~dlum of th- pl~t~l~t lnv~nt~on, but ~o~ not ~ont~in dextro~o ~nd hA~ A lowor ~odlum chlorld~ cono-ntratlon of 5.23 gr~m~ p~r litor. ~ho symbol ~M ~ dextro--~ r-pr-~-ntr a otor~g~ ~odlu~ h~vlng tho ~ ch~cto~tlc~ of th- proforr-d pl~tol-t torag- m-dlum of th- plat~l~t lnvontlon, lncludlng doxtro~, but h~ th~ lowor ~odiu~ chlorld -33 1326~ conc~ntr~tlon o~ 5.23 gr~m~ p~r llte~. ~he ~ymbol "DMSM" repr~sent~ a ~tor~ge medlum havlng the ~ char~cterlst~c~ of the prefereed platele~ ~torage m~dlum, but do~s not contaln dextro~e, The comp~r~tlve example~ do not repr~s~nt the ln~entlon, TAa~ 13 D~y l D~lsr 5 DAy ~ O C.

Ex . E BSM 66 ~ 8 47 + 11 C.

Ex . G BSM ~ d~xtro8e~3 ~ 8 73 + 10 C.Ex. 8 DM8M 8~ +10 56 1 9 ~x. 6 P.S.M. 95 + ~ 91 1 3 TA~L~ lq NyQotonlc 8hock R--pon~ R~cov~y~ Day 1 D~y 5 D~ O C.Ex. F B~M 28 ~ 5 20 ~ 6 4 ~ 10 C.~x. G B8M + d-xtroJ- 44 ~ 10 44 ~ 1022 1 lO C.~x. H DM8M 7~ ~ 13 41 1 lO~ 1 5 ~x. 6 P.8.M. 73 ~ 20 61 ~ 1351 1 6 -3~ ~ `J 132~0 TA~r~lS 1 5 .

~, DAY SD~Y 10 C.Ex. F BSM l.~ I i 7 ~ 3 2 ~ 2 C .

Ex . G BSM ~ dextroae 16 ~ 3 10 1 4 5 t 3 C.13x. H DMSM 15 ~ 3t ~ 2 1 _ 1 Ex. 6 P.8.M. 17 ~ 414 ~ 2 10 _ 3 TABLE 1 6 Rato of Ox~on con~ulQption nmol~s~ n/10~ pltB~ ~L ~,D~y 5 D~y 10 C.Ex. ~ BBI~ 1.0 t~ 0.1 0.5 + 0.3 0.3 ~ 0.1 C.15X. ~ BSM ~ d~xtro~o 1.0 + 0.3 0.6 ~ 0.1 0.6 _ 0.~ C.ISx. H DMSM 0.9 + 0.3 0.6 ~ 0.1 0.2 1 0.1 Ex. 6 P.S.M. 1.0 + 0.1 0.7 -~ 0.1 0.4 0,3 TJ~BLII 17 TP~ nmol~l09,~plt-~ D~Y- I Do.~c 5 D~y 10 C.15x- H DM8M 7.1 ~ 1.5 5.3 1 2.2 0.7 1 0.2 ~:x. ~ P~M 8.5 ~ 1.5 ~.4 ~ 1,3 6.5 + 0.

_ _ ~R--ult- r~pr-~-nt m-an ~ tand~rd dov1lltion.

_~5_ `- - / v 13264~ ~ he result~ o thl~ oxa~pl~ and ~omparatlv~ OX~mP1e6 d~m~n~tratO thO fO11OW1ng~ ~ 1) A m1n1mU~ Of 3000 tO 6000 m~ gr~mB Per 11ter Of 60d1U~ C1trate Wa8 e~nt1~1 tO a~Oid P1~O1Ot C1UmPin9 and ~UbgOqUQnt dOtRr1OrAtiOn Of th~ P~AtO1Ot6~ ~ 2) GOOd malnt~n~ne~ of platelo~ di~oid ~orphology and neg11g1b1e C1UmPin9 Wat Obtain~d W~th ~Od1~ Ch1Or1dO 1n the COnCentrat1On r~n9~ Of 64000 tO 76000 m~ grAmB P~r 11tOr. Poor quallty o~ the P1Ate1et COnCOntratO WA# O~erYed 1n the data O~ Tab~ W1th 8Od1Um Ch1Or1dO R~ a COnCentrAt1On Of 5.23 gram~ POr m~ ter, t3) ~he add1t1On O~ d1V~1~n~ Oa~1On~ ~Mg~ and Ca~) W~4 neO~rrarY fOr ma1ntOnan~- O~ P1~tO1et dl~cold mOrPhO1O~Y. CA1O1U~ chlorldo can bo us-d ln the COnCentratiOn r~nge o 100 tO 400 ~ gram- P-r 11t-r and magne~lum ohlorld- can b- u~-d ln th- ran9~ Of 200 to 400 mllllgr~mr por 1Iter W1th good rO-U1t8.

~ 4) ~ota~alum chlo'rli- w~ ntlal to ~Alntaln ` no~al morphology and war u--d ln th- COnCOntrat1On ran9~ O~ 220 tO 400 m~ 9ra~r POr 11t-r wlth good ~8U1t~.

~ 5) 80dlum dlpho~phat- WAB u~od ln tho conc~ntra~lon r~nq- of 100 tO S80 m111i9rA~r P-r 11t-r wlth good rO~U1t~ --36- 1326~0 (~) sodi~m hic~rbonat~ w~s u~ed in the tho concentration ranqe o~ 2~00 to 4200 milligram~ per litsr with good results. A mlnlmum of 2~0 milligra~ per liter w~ feund to be neo~ ry in ord~r to prevent a decre~8e in pH w~ th stor~ge be~ond ~even d~y~.

~ 7) A 2.5 to 7.5 percent carbon dioxlde atmo~phe~e, ~ependln~ on the amount o~ sodlum blc~rbon~te ~d~ed, wa~ ~ucc~fully used ~ maint~in ~ ~on~t~nt pH.

~ 8) ~he a~dition of gl~co6e (dext~o~e) wa~ essential for m~lntenAnce of good in vitro quality of the pl~telets a~ evld~nced by the dat~ of ~bl~ 13.

Dextrose wag u6ed ln the conoentr~tlon r~nqe Of 3000 to 7500 milllgram~ per llter wlth good ~e~ults.

~ he p~ of the pl~telet ~tor~ge modiu~ wa6 mAlnt~lned ln the re~nge of 6 . B to 7,4 ~: me~c~red at 22C. A p~ above 7.4 resulted ln cl~mpin~ o~ the pl~t01ets while a pH below 6.8 cAused swelling ~nd 10~6 Of dl~oold morphology.

EXAMPLE ? _ AND COMPARA~ = The slmllarl~y ln chemleAl ~nd phy~iological attri butes o~ the pl~telet GtOrige medium to CP~-plasma are indicative of the inherent non-toxic and safe utility characteristics of the platelet or red blood cell ,.~,i~ 13264~ ~tor~o al~dlu~ ~or ln~uslon lnSo patlent~ lono~ ln l~o ~tudy c platolot~ pr~rved wlth tho plotolot ~orogo ~dlum o~ thl~ ~nvontlon w~e conduct~d.

The BtU~ aa~cr~b~d hereln onco~p~so~ 10 palro~ ~tudl~s.

The p~lrod 4~udlo~ u~lng notmal 11101~16 ov~r tho age of 21 not known to hove ~ontal or phy~lc~ abillty ~n~l not rooolvln~ dru~ th~r~w.

Tho vol~ntosrs don~tod ono unlt o~ pl~t~lot rloh plos}Q~ whlch wa~ drawn u~lng tho oonvontlonol-platol~t ~phar~ls t~chnlquo~, Sh~ ~eooodueo wae p~r~or~od twloe. ~he pl~olot conc~ntr~t~ wor prooo~o~ ~oe etor~g~ for ~ 7 day p~rlod ot 22C olthor ln CPD-P1~mO Or ln tho roa c~ tor~go ~o~lum u~lng currontly llcon60d procodue- Pl~tolo~ ooncontr~to- woro ~torcd ln ~n ~glt~t4r~1ncubatoe at about 22c. The platolet aono~ntr~t-~ ln ~PP-P~ w-ro ctor-d ln ~n ordlnary ~1F atmosphoro sho pl~tolct conc-ntr~tor In tho pl~tolo~ ~torogo ~odlu~ woro 8tor-d und~r ~ 7.5 p~rcent C2 atmo~ph-r-, Thl~ jeoooarlng and ~torag- o~ pl~tclot ooncontr~to~ ln C~D ~l~o~ conrl~tent ~lth thourr~ntly llconrod proc-~ure Tho ln vlvo ~l~blll~y o~ th- platolot~ w~dotor~lnod by tho oonv~ntlon~l p-rcont rocovory An~ ~urvlval par~o~or~ urlng r~lol~otoplo labollng toohnlquor woll known ln tho ~rt ~uoh ~ dlrou-o~i ln -3~ v '~ 132~ "Platele~ Klndtlcs ~nd sm~ging", Volume I, Technlque~ ~nd ~ormAl PlAtelet ~lnetlc~ yn~ et a1., CRC Pr~ Inc., Boca Raton, Florld~ ~1985),.

At the completlon o~ 7 d~y~ o~ 8tor~ge, lO mlllllltars of pl~telot concentrate wer0 t~ken o~ radlol~otoplc lnbelln~ of tb~ pl~t~lets wlth 111 Indlum-Oxln~. The wa~hed and l~beled platelet4 wore rQsu~pend~d ln 6 mlllil~te~ o~ nonr~dioaotl~e autologo~s pl~sm~ ~or infu6~0n into the orlglnal dono~.

Two mllllllt~rs o~ blood ~amplos were dr~wn from the donor~ at l, 2, and 3 hour lnterv~ls aft~r lnfuslon ~d then dAlly there~fter for 7 d~y~ for c~lculatlon of ln vlvo percent recovery ~nd ~ucvlval. The ~tudy wa~ de41gned 6uch that durlng the fir~t 4e~10n S ~onor~ wcre ln~u~e~ w~th plAtel-ts ~tored ~n th~ platel~t storAge m~dlum o~ thl~ lnv~n~lon and 5 donor4 wor~ lnfused wl~h plAt~let~ 4tore~ ln CPD~plasmA. ~h~6 w~a r-p-Ated durlng th~ ~econd ~ lon, 2 month- later, wlth th- ~torAge medlu~ b~lng rov~t#~d ~or the donor8. Tb~ ~erc-nt re~ov~rles and ~urvlvAl- w-r- dQt~rmln~d u~1ng the g~mma ~unctlon multlple hlt program. Palrod t-t-st~ w~re u~-d to d~t-c~ ~lgnl~lc~nt dl~-renc-~. Tho ln vltro vl~blllty o~ ~ho platelet~ wn~ ~valu~tcd by -39 ~326~0 hypotonlc shock re~ponæe And ~xtont of ~hap~ oh~ngo wlth ADP.

The r~ul t~ o~ the peroent o~ ~n vLvo recovery ~nd surv~v~l6 Are ~hown in ~bl~ 18 ~nd 19, ro~pcctively.

~e~!!

I VD~ .Ex, ~ Ex. 7 ~onor ~Asmzl P.lS.M.

29 57 2 23 52 3 36 5~ 43 47 31 S~ 6 61 66 7 3~ 48 8 28 37 9 41 44 0 43 . 55 m-An + t~nd~rd ~lo~ tlorl 37 ~ 11 51 ~ 8 -40 ~32~0 'T~EIL~_~ C.~5x. I 15x. 7 Donor ~e~ P,#.IS.

54 125 2 85 162 3 126 159 ~1 17~. 145 103 155 6 86 146 7 12a 150 8 ~07 114 9 129 154 1o 121 130 m-~n ~tand~rd a~vlatlon110 ~ 32 144 + 16 Moon poroont ln v~o raco~orlos and ~urvlval~ woro found to bo 8ubstAnti~lly hlgh~r wlth ulat~l-t conc~ntr~tos ~torod ln r~d c~ eor~go ~cdlum or 51 + 8 p~rc~nt and 144 + 16 hour~ vor~u~ ~7 ~ 11 p-ra~nt ~nd 110 _ 32 hour~ ~oe platel~t concontrAt- storod ln C~D-pl~4~ o~p-ctlv-ly. ~ho dl~or-no-~ w~o 4tatl~10~11y, hlghly lgnl~lcant ar wltn-440d by --4 1- ~J ~ j;

~32~4~0 t-te6t v~lue of P c O.OOS.

Ths ln vltro v~411~ty re6~1ts pAr~lleloâ Sho n vivo r-rult~ as evldcnced by the data pr~ent,ed in Tablc~ 20 ~nd 21 wlth 6t~t1~tlo~11y 6~perlo~ re~ults indlcot~d by the pAlrod t-te~t v~lue of p<0 . ûl for pl~telet conc~ntr~te~ ~tor~d n pl~tolet 6tor~ge medium A~LS 2 0 HYDotonic Shock ~csponseC_ 96 RocoverY C.Ex. 1 Ex. 7 onor CPP-pl~6m~P.8.1t.

18 75 2 Is ~7 3 4~ 62 4 40 ~0 ~5 ~8 44 70 7 4,7, 67 8 40 S0 g 50 55 lû 77 lO0 m-an r t~nd~ rd v1ation 44 1 15 65 ~ 14 --42- ~J . ~_J 13264~0 TA~L~ a C.~x. I ~x. 7 ~onor CPD-PlA~ma P.~-M.

__ 1 4 la 2 6 1-~ 3 ~ 19 4 7 14 g 10 13 6 8 14 7 1? 18 a 7 11 g 14 11 0 19 17 ~n ~ ~tandard aevl~tlon 9 ~ 4 14 ~ 3 Th0 r~sult~ o~ thl~.~xAmplo And ¢ompA~Ativ~ ex~plo lndlcat~ th~t th~ ln v~vo vlablllt~ o~ pl~t-l-t conc~ntr~t~ ~ro ~ubstantlally lmpro~d ln th- pl~t~lot stor~go modlum o~ thi- lnventlon when co~par~d to Rl~tolot ~torAg~ ln C~D-pl~ma.

-43 ~L326~ 0 ~ ~ EX~MPLEE; B ~ OUGN 15.1U~D COMPARATIVIS I~XAMPLI!: J ~ ho prooodure uoed ln thq~ ~x~ploB ~nd thl~.

comp~cJ~tlvo ~x~plo ~o ~opar~t~ ~od blood co11- tro~ whole blo*d and ~o m~k~ tho otor~ dlu~ ~r~ the ~m~ tho~ de60rlbod ~bov~. lSxAmplo~ ~ through lS ~re ~lrecto~ to th~ rod blood coll ~or~ dlu~ of tnc ~n-,rentlon. CO~PA~t1VO ~x~mplo J d~on~r~tos the curr~nt BtAte of th~ art ln r~d blood c~ tora~e modlu~8, ~o~p~r~tlvB ExA~ple J uses th~ r~d blood cell stor~g~ modlum ~old. un~r the ~r~ oL~9.

~ x~mpl~ 8 throu~h 15 And co~pAr~tl~e ~xJ~pl~ J woro p~rforlnod wlth 10 wholo blood unltr collecte~ lnto CPD ~tloo~gul~nt ~nd dl~ldod ~ tlcally lnto ~our 6u~unlt~ to obt~ln 40 tot~l to~t unltr, ~rho blood was ooll~ct-~ ln 150 ~lllllltOt ~-n~al PL-Iq6 plaBtlc tr~n~fet pAck~.

Aft-r th- blood wa~ o-ntrl~ugod, a ~u~tl¢l-nt guantlSy of a ~llt-rod, st-rll- !tor~g~ ~dlum wat ln~ootod to prod~oo ~ flnal h~atocrlt ~f 60 S p~ro-nt.

Exa~pl- 8 1- dlr-ot~d to th- ~tora p of r-d blood a~llB wlth th~ r~ bloo~ c~ll otorAg~ ~dlu~ of th~ ln~ntlon h~vlnq ~ low gluco-~ ~LC) aoncont~tlon o~ 7 grA~ of d~xtro~c p~r lltor.

exa~pl~ 9 lr dl,rooSod to tho ~tor~g~ of rod blood ooll~ wlth tho r-~ bloo~ c~ torAg- ~odlu~ ot thl~ ~ v 1326~0 lnven~lon h~vlng a hlgh gluoo~e IHG) conc~ntrAtlon o~ 20 gram~ o~ dextro~e por liter.

Ex~plo ~0 16 di~ectod to the ~torage o~ red blood cell~ wi~h the red blood cell ~tor~g~ ~edlu~ of thl~ lnvention havlng th~ low gluco~e concentratlon o~ Ex~mple B and 2.5 ~raas (30mM) of 40dlum bloa~bonate ~G~ ex~ple 11 13 dlrected to the ~torage o~ red blood c~118 with ~he red blood cell storags m~dlum of thl4 lnventlon h~v~nq tho hlgh glu0080 ~oncentratlon o~ ~x~ple g and 2.5 grAms t3~mM) o~ ro~lu~ blcarbon~t~ (~G~B).

~ xamplo 12 l8 d~rected to tho ~torage o~ red blood cell6 wlth tho red blood coll ~tor~ge medium of thl6 lnvontlon h~vlng the low gluco~e ~onc~ntr~tlon of Exampl~ 8 ~nd 270 mllllgeams of ~do~lno por llter t L~ ~ x~plo 13 1~ dlroctod to tho ~torago o~ r~d blood oolls wlth thc rcd ~lood coll storag- ~-dlum o~ thlln~ntlon h~vlng tho hlgh gluco~o conc-ntratlon o~ ~xa~pl~ 9 and ~70 mllllgr~ o~ ad-nln- p-r llt-r Ha~ ~ xamplo 14 1~ d~octoa to tho torag~ o~ r-d ~lood ooll~ wlth th~ rod blood ccll atorag- ~od~um o~ thl~ ln~-ntlon having th- low gluco-e conc-ntr~tlon or -45 1326~0 `~' Exampls 8 wlth a. s grAm~ o~ ~odium bic~rbonate and 270 mllllgram~ of adenlne pcr ll~er ~G/~A).

~ x~mplo 15 i~ ected to the ~orag~ of red blood c0116 wlth the red blood cell 6torage medlum of th~ ln~ntion hav~ng the hlgh gl~co~e con¢entr~tlon of Ex~mple 9 with ~ ~ gram~ o ~od~u~ bicarbonate and 270 mllllgram~ of Adenlne p~F lit~r (HG/B~A) Thl~ repre6ents the pr~ferred embodlment of the red blood cell ~oraqe medlum Comparatlve ~xample J 18 dlrected to th~ ~torag~ o~ r~d blood c~ wlth the r~d blood cell storage medlum ~old und~r tho mark ADSO ~ whlch ha- 9 grAms of ~odlu~ chlorld-, 7 5 gram~ of mannltol, 22 gram- of d-x~ro~, and 270 mllllgra~- of ad~nlne per llt-r Samples of ~ mllllllt~rs each of tor~d r-d blood cell- w-r- drawn ln ach o~ th- ~bov- ex~mplo~ ~nd comparAtlva ~xampl~ for blocho~lc~ y ~ho-o rampl-~ w~r- drawn lmm~diAtely a~t~r b~lng peo~arod with th~ r--p~ctlv- torag- roiutlonr ~nd on day~ 7, 14, 21, 35, and ~9 of torAg- at ~C ~h- blochomlcal ~6~yB wer- ~or d-termlnatlons of th- p-rc-nt lo~ o~ ASP tho p~rc-nt h-moly~l~, th~ pH ln olutlon, and tho p~rcent lo~r of 2,3 DPG of tho tor~d r-d blood c~llB~ Flgur~- 1, 2, 3, and 4 graphlcally doplctB tho d~ta o~ tho rour bloch-mlcal a~-ay d-rcrlbod ~bo~ tor tho 132~4~0 49 d~y storage period. ~he~e ~lguree preeent tho me~n~, plus or minus a standard deviation, of five studies wlth each of the eight embodiment~ of the red blood cell s~orage med1um u~ed in Example~ 8 through 15. ~he~e data are compared to me~n r~8ult8 of 6 subunlt0 o~ blood stored in the A~S0 ~ brand storage medlum.

The re~ult6 of the~e example~ and aompar~tlve ex~mple a~ lllustrated by Flgures 1 and 2 demon8trate that red blood cells stored ln the storage medlum6 fortl1ed wlth 60dium blcarbonate and ~denlne h~d superlor ATP malntenance ~nd les~ hemoly61s after 4~ d~y6 than the control red blood cell~ thAt were etored ln the ~DSO~ brand stor~ge medlum. suprl4ln~ly~ good ATP level8 were achleved wlth the red blood cell 5~0r~ge medlum of thls lnventlon not h~vlng adenlne. ~he re6ults obeerved for these ex~mples were ~llghtly le~5 than the results observed for the ADS0 ~ medlum.

The resulte of the6e ex~mple6 and comparative example a~ illu~trated by Figure~ 1 and 2 demonstr~te that red blood cells stored ln the storage mod~um fortlfied wlth ~odlum bic~rbonate have desleabl~ high pH level~ and 2,3 DeG level~. No Bl9nlllcant dlS~erence waB ob~erved ~etween red blood cell~ ~tored ln the storage medlum of thla lnventlon wlth elther low oe nlgh , ~ 1 3 2 ~ O glucobe con~entr~tlon~ when compared to red blood o~ storoa in the ADso ~ brand storage madium Th~6e examples and comparatlve example demon~trats th~t the red blood c~ torage medium of thls lnventlon ~ortifled wlth both ~odlum bicarbon~te and Adenlne produce~ de4lrable A~P malntenance durlng ~torage XA~PLX 16 AND COMP~kATIV0 EXAMPLE8 X THORUGH o ~ he procedure u~ed ln ~xomple 16 to ~p~r~te red blood coll~ from whole blood and to mako the ~torage ~edlum are th~ same a8 tho~e d-rcrlb-d above, but wlth a dextro~e oonoentr~tlon of 7 0 grams per llter and ~ ~odlum blcarbonate conc~nteatlon of 3 gra~s pee l~ter Comp~r~tlve Exampl-~ ~ through o w~r- not p~rformed by th~ lnventor6~ laboratory, but r~pre~ent dat~ reported by oth~rs ~ h- data of thlr xampl- and ~omparatlve ex~mpl~ ~re d~rl~ed from te~ts performed on whole blood unlt~ f~om 5 donor~ th~t w~r~ oollocted ~nd proc~ d aooordlng to ~tand~ed blood bank m~thod~ ~n tho ~x~mpl- tho r-d blood cell~ w-re susp~nded ln l00 ~llllt-r~ of the pr-~orr-d embodlm-nt of th- platelot ~torage m-dlum h~vln~ A concentratlon o~ 270 m~ gr~m~ per llt~r or ~denlne Tho ~uspendca rcd blood c~ wero 4tored ln 600 mllllllter Fenw~l P~-146 prlm~ry 13254~0 `W~ pl~st~ c bags at ~~ ~or up to 56 d~yfi .

The re~ult~ s~b~rvod by thl~ ~xnmple And comp~r~tlv~ cxAmple6 arc preee:~ted ~ n Tablel~ 22 ~nd 23, _~9_ ~ - " ~3264~ t ~D N o ~ q~ 0 ~ . ,., " g g ~ 8 m ~ ~ ~ ~a o ~ ~o ~u O ~D ~ `4 ~ 't ~ o~ o o o " .

~ ~ U'~ o c o o C~ oc ~U ~ o o N ~1 ~ ~J ~ g~ 1~1 ~ ~ O ~ ~ ~ C~ ~ C ~ ~: O U~ U~ ~ ~ U'~ o ~ ~ ~ ~ ~ V ~ ~ ~ 8 ~ ~ o o !l 3 ~ e ~4 ~ ~ ~ o v t~ so v 13~64~ -1 N O O C ~ ~ o 0 ~0 ~0,~1 N ~ ~1 ~ O I t~ o C> o ~ ~ 1~ 0 .~ ~ ~-3 O O O - ~ ~ ~ ~:

r~ r --I N 8 N¦ ~ ~¦ O O c ~ a 51 W Wl ~ ,~ O O ~ X ~ ~ 0 g ~ X Q ~ o ~ ~ ;~ O a, .~ . . .

V C~ ~ V --51 ~ - , 1326~0 The re~ult~ of thls exAmple and these comp~r~tlve ex~mple~ demon~trate th~t the red blood cell ~tor~ge medium of th;~ lnvent~on pro~lde~ more de~r~ble A~P levele And less hemolyBl~ in red blood cells stored or more than 49 days th~n ob~e~ved in red blood c~116 ~tored in other red blood cell ~torage mediums.

EixAM pL~s 17 Tho pr~cedure~ u~ed ln Ex~mple 17 to ~p~rate r~d blood cell~ from whole blood and to make the ~torage medium i~ the ~me a~ those described above.

The ~tudies des~rlbed a~ for Example 16 used the platelet ~torage medium wlth 3 grams per l~ter of sodlum blcarbon~te ~nd PL-146 pla~tlc contalner6. This rasulted ln a decrea~e ln pH of th~ red blood cell solutlon to below 6.6 after 28 day~ o~ 6toraqe.

In Ex~mple 17 red bloed cells ~r~ stored accordlng to the procedure Or 6xample 16, but ~erumo X~-612TM platelet contalner$ made of thln polyvlnylchlorlde ~PV~) plA~tlc and dlethylhexylphth~late ~EHP) are used ln6tead of PL-146 cont~iners. Addltlon~lly, thC ~od blood oell6 are stored wlth a 50 porcont ~oductlon ln thickness when compared to the thlcknes6 o Examplo 16 and wlth an lncrea~ed ~odium blc~rbonate concentratlon of 4 grAm6 per llter. The red blood cell volume~ u~ed ?

1326~ l' 0 in the 400 milliliter Terumo XT-612 M containers are two-thlrd~ o~ the orlglnal volume, This i6 to provlde the ~me r~ti~ ~f red hlood cell volume to eont~iner volume a~ u~ed for t}le 600 milliliter PL-146 contain~r~ ef Ex~mpls 16.

F1ve ~tudles w~re performed to di~eover whether or not the lncr~a~ed Co2 permeabllity oP th~ ~erumoTM cont~iner~ resulted ln h~qh pH level~ and improvsd red blood cell qualitie~ a~ter ~torage. The 6.4~ + 0.08 pH ~ the red blood cell~ stor~d ln the Ter~mo XT-~l~TM cont~iner~ wa~ only 61ightly higher than the 6.39 0.09 pll of the red blood cell~ 6tored in the P~.-146 cont~iner~. Surprl~lngly, the red blo~d cells stored in the TerumoTM containers had improved ATP levels ~nd le~ hemolysl~ durlng 6tor~ge than did the red blood cell6 6tored in the PL-1~6 contalners. After 56 days o~ Bt~r~ge the m~n Arp level was 2.54 + 0.57 umoles/gHb ~nd hemolysi~ wP8 0.38 + 19 percent. Thl6 0x~mple dcmon~trate~ that prolonged ~torage o~ red blood cells ln the storage medlum o~ thl~ ln~ention i~ ~uperlor to the results obtalned with other, commerclally avallable pre6erv~tion ~olutlons. Furthermore, this example demon~tr~te8 th~t plateletr and red blood ~ell~ can be Gtored in the ~me type of plaEtlc cont~iners.

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