Coagulant stability and sterility of thawed S/D-treated plasma

BACKGROUND: Units of frozen S/D-treated plasma (SDP) must be transfused within 24 hours after thawing. To avoid waste, an attempt was made to determine how long SDP could be therapeutically effective after thawing and storing it at 20 degrees C. STUDY DESIGN AND METHODS: The microbiologic safety and the activity of labile coagulation factors were evaluated in units stored at 20 degrees C of thawed SDP units and FFP within 24 hours of collection (FFP24). Five SDP and FFP24 samples of each ABO blood group were cultured and assayed for coagulation factors daily over 5 days. Assays included FV, FVII, FVIIa, FVIII, F IX, FXI, protein S, antiplasmin, fibrinogen, prothrombin times (PTs), and activated partial thromboplastin times (aPTTs). RESULTS: None of the 80 bacterial cultures demonstrated growth under either aerobic or anaerobic conditions. FV, FVIII, F IX, FXI, fibrinogen, and the aPTT appeared to be stable in both thawed FFP24 and SDP. The PT increased slightly in thawed FFP24 and insignificantly in SDP. FVII decreased slightly in FFP24 but remained in the normal range, and FVIIa was low and constant. FVII was increased in SDP and FVIIa was markedly increased. Protein S decreased from initial normal values in FFP24 to very low values. Protein S was very low immediately after thawing in the SDP and continued to decline. Antiplasmin was normal and stable in thawed FFP24 but was low in SDP and remained constant after thawing. CONCLUSION: Sterile SDP that is stored at 20 degrees C provides sufficient coagulant activity of labile FV and FVIII to transfuse it for up to 5 days after thaw. Caution is warranted by decreases in Protein S and antiplasmin, clinical evidence of coagulopathy in some recipients of SDP, and a recent manufacturer's warning.     

Storage of thawed plasma for a liquid plasma bank: impact of temperature and methylene blue pathogen inactivation

BACKGROUND: Rapid transfusion of fresh‐frozen plasma (FFP) is desired for treating coagulopathies, but thawing and issuing of FFP takes more than 40 minutes. Liquid storage of plasma is a potential solution but uncertainties exist regarding clotting factor stability. We assessed different storage conditions of thawed FFP and plasma treated by methylene blue plus light (MB/light) for pathogen inactivation. STUDY DESIGN AND METHODS: Fifty thawed apheresis plasma samples (approx. 750 mL) were divided into three subunits and either stored for 7 days at 4°C, at room temperature (RT), and at 4°C after MB/light treatment. Clotting factor activities (Factor [F] II, FV, FVII through FXIII, fibrinogen, antithrombin, von Willebrand factor antigen, Protein C and S) were assessed after thawing and on Days 3, 5, and 7. Changes were classified as “minor” (activities within the reference range) and “major” (activities outside the reference range). RESULTS: FFP storage at 4°C revealed major changes for FVIII (median [range], 56% [33%‐114%]) and Protein S (51% [20%‐88%]). Changes were more pronounced when plasma was stored at RT (FVIII, 59% [37%‐123%]; FVII, 69% [42%‐125%]; Protein S, 20% [10%‐35%]). MB/light treatment of thawed FFP resulted in minor changes. However, further storage for 7 days at 4°C revealed major decreases for FVIII (47% [12%‐91%]) and Protein S (49% [18%‐95%]) and increases for FVII (150% [48%‐285%]) and FX (126% [62%‐206%]). CONCLUSION: Storage of liquid plasma at 4°C for 7 days is feasible for FFP as is MB/light treatment of thawed plasma. In contrast, storage of thawed plasma for 7 days at RT or after MB/light treatment at 4°C affects clotting factor stability substantially and is not recommended.     

Analysis of prolonged storage on coagulation Factor (F)V,FVII, and FVIII in thawed plasma: is it time to extend the expiration date beyond 5 days?

BACKGROUND: According to AABB standards, fresh‐frozen plasma (FFP) should be thawed at 30 to 37°C and expire after 24 hours. An increase in the aggressive management of trauma patients with thawed plasma has heightened the risk of plasma waste. One way to reduce plasma waste is to extend its shelf life, given that the full range of therapeutic efficacy is maintained. We evaluated the effect of prolonged storage at 1 to 6°C on the activity of Factor (F)V, FVII, and FVIII in plasma thawed at 37 or 45°C. STUDY DESIGN AND METHODS: Group O plasma from healthy donors (n = 20) was divided into 10 pairs and frozen and stored at not more than ?18°C. One sample from each pair was thawed at 37 or 45°C, and all were stored at 1 to 6°C. Samples were analyzed for FV, FVII, and FVIII activity on Days 0, 5, 10, 15, and 20. RESULTS: Plasma thawing time was 17% less at 45°C than at 37°C. No differences were observed between thawing groups in coagulation activity of FV, FVII, and FVIII during the 20‐day storage period (p > 0.12). In both groups, the activity of FV and FVIII decreased over time but remained within a normal range at 10 days. CONCLUSION: Although levels of plasma clotting factors are reduced in storage, therapeutic levels of FV and FVIII are maintained in thawed plasma stored for up to 10 days at 1 to 6°C. Thawing of FFP at 45°C decreases thawing time but does not affect the activity of FV, FVII, and FVIII.     

Evaluation and comparison of coagulation factor activity in fresh-frozen plasma and 24-hour plasma at thaw and after 120 hours of 1 to 6°C storage

BACKGROUND: Current transfusion-related acute lung injury reduction strategies include avoiding transfusion of plasma products collected from female donors or female donors that have been pregnant to reduce transfusion of plasma-containing HLA antibodies. Such a policy considerably decreases the number of donors available for generation of fresh-frozen plasma (FFP). To increase the supply of FFP, substitution of 24-hour plasma (FP24) and thawed plasma (TP) derived from either FFP or FP24 may be viable substitutes. To justify such a policy the coagulation factor content of FFP, FP24, and TP derived from both product types was assessed.
STUDY DESIGN AND METHODS: Coagulation factor (F)II, FV, FVII, FVIII, F IX, and FX; protein C (PC) and protein S (PS); von Willebrand factor antigen and ristocetin cofactor; fibrinogen; and antithrombin activities were analyzed in nonpaired FFP and FP24 at the time of product thaw and again after 120 hours of 1 to 6°C storage.
RESULTS: At thaw, mean FVIII and PC activities were lower in FP24 than FFP. Mean PC and PS activities were lower in FP24- than FFP-derived 120-hour-old TP. No other differences in mean activity reached significance. Activity levels were generally lower in TP; FVIII, FV, and FVII showed the largest changes. However, prestorage leukoreduction appears to improve the stability of FV.
CONCLUSION: FFP, FP24, and the derived TP all contain adequate coagulation factor activities to maintain hemostatic activity. As FFP becomes less available, increased use of FP24 and TP are viable alternatives for most clinical situations.     

Activity of clotting factors in fresh-frozen plasma during storage at 4°C over 6 days

BACKGROUND: Fresh-frozen plasma (FFP) requires thawing, which delays availability. We investigated clotting factor activity and bacterial contamination of FFP when stored at 4°C ± 2°C for 6 days.
STUDY DESIGN AND METHODS: Plasma of 20 healthy plasma donors was sampled, frozen, and analyzed at baseline and repeatedly over a period of 6 days after thawing. The activity of fibrinogen, Factor (F)II, FV, FVII, FVIII, F IX, FX, XI, FXII, FXIII, antithrombin III (ATIII), von Willebrand factor antigen (VWF-Ag), protein C (PC), and free protein S (FPS) were determined and analyzed over time.
RESULTS: Immediately after thawing there was a significant decrease of fibrinogen (−9%), FII (−7%), FV (−14%), FVII (−12%), FX (−11%), FXIII (−20%), PC (−7%), and ATIII (−4%), whereas FVIII (+8%), F IX (+1%), FXI (+11%), FXII (−1%), FPS (−1%), and VWF-Ag (−6%) remained stable without significant change. Over 6 days after thawing fibrinogen, ATIII (+2%) and VWF-Ag (+2%) remained stable whereas FXII (+2%), FXIII (+6%), and PC (+3%) changed significantly over time and increased at the end. FII (−8%), FV (−16%), FVII (−31%), FVIII (−47%), F IX (−12%), FX (−10%), FXI (−25%), and FPS (±0%) changed also significantly over time and decreased at the end. All clotting factors and inhibitors remained within the reference range requested by quality assurance regulations. No FFP bag showed bacterial contamination.
CONCLUSION: This provides evidence for maintaining quality of thawed FFP and may improve rapid availability in emergency situations and reduce cost for health care givers.     

Postfiltration factor VIII and fibrinogen levels in cryoprecipitate stored at room temperature and at 1 to 6 degrees C

The 13th edition of the standards of the American Association of Blood Banks specified storage at 1 to 6 degrees C for cryoprecipitated anti-hemophilic factor (Cryo) administered up to 6 hours after thawing if the Cryo is used for factor VIII (FVIII) content (Standard J4.210). Previous editions specified room-temperature (RT) storage for up to 6 hours. Currently, the temperature specification has been deleted. There are few data addressing the optimal storage temperature and maximum storage time for FVIII and fibrinogen in thawed Cryo. Thirty bags of Cryo were assayed for FVIII and fibrinogen. Each bag was divided into two aliquots; one was stored at RT and the other at 1 to 6 degrees C. Assays were performed immediately after thawing (Base) and 6 and 24 hours after thawing, respectively. All samples were filtered through 200-mu blood component infusion sets before assay. Three hundred analyses were performed, 150 each for FVIII and fibrinogen by conventional clotting technique. Data were analyzed by using a paired t test. Cryo stored at 1 to 6 degrees C for 6 and 24 hours showed an FVIII loss of 35 percent (p less than 0.0001) and 63 percent (p less than 0.0001), respectively. Cryo stored at RT for 6 and 24 hours had an FVIII loss of 8 percent (p greater than 0.05) and 20 percent (p less than 0.0001). Cryo stored at 1 to 6 degrees C for 6 and 24 hours had a fibrinogen loss of 20 percent (p less than 0.0001) and 43 percent (p less than 0.0001). Cryo stored at RT for 6 hours had no fibrinogen loss and a 2 percent loss at 24 hours (p greater than 0.05). These preliminary data show a significant loss of FVIII and fibrinogen activity in Cryo stored at 1 to 6 degrees C and filtered before assay. The FVIII and fibrinogen activity at RT is clearly maintained up to 6 hours after thawing.     

Stability of solvent/detergent-treated plasma and single-donor fresh-frozen plasma during 48 h after thawing.

The aim of this study was to compare the quality of solvent/detergent (SD) treated plasma, Octaplas, and single-donor fresh-frozen plasma (FFP) units during 48-h storage after thawing. Octaplas bags of different blood groups and individual FFP units were thawed and stored at either +4 degrees C or at room temperature (RT) for 48 h. Samples drawn during the observation period were investigated on various coagulation factor and protease inhibitor activities using standard coagulation and chromogenic assays. The generation of FVIIa was followed as a marker of coagulation factor activation. All investigated coagulation factors and protease inhibitors were stable for at least 8h during storage of Octaplas at +4 degrees C. FVIII levels started to decline earlier in FFP than in Octaplas at both storage temperatures. Stored Octaplas OD660 values were more stable during the storage period than FFP OD660 values, whereas VWF multimeric patterns were comparably stable in both types of plasma. In conclusion, this stability study has demonstrated that thawed Octaplas maintains its high quality, even with a time safety margin, for 8 h at +4 degrees C and for 6 h at RT. In general, there was more variability in coagulation factor levels among different FFP units compared with different Octaplas batches.     

Quantitative and qualitative analysis of coagulation factors in cryoprecipitate prepared from fresh‐frozen plasma inactivated with amotosalen and ultraviolet A light

BACKGROUND: There were no previous studies about the quality of cryoprecipitate prepared from fresh‐frozen plasma (FFP) inactivated with amotosalen and ultraviolet A (UVA) light. The aim of this study was to analyze the quantity and quality of coagulation factors in cryoprecipitate prepared from FFP treated with amotosalen and UVA light. STUDY DESIGN AND METHODS: FFP was obtained from whole blood donations and inactivated with amotosalen and UVA light according to the manufacturer's instructions. Fibrinogen, factor VIII (FVIII), von Willebrand factor antigen (VWF : Ag) and activity (VWF : RCo), the von Willebrand factor cleavage protease activity (ADAMTS‐13), and the multimeric structure of VWF were analyzed. RESULTS: The content of fibrinogen, FVIII, and ADAMTS‐13 was lower in cryoprecipitates prepared from amotosalen‐treated plasma when compared with cryoprecipitates prepared from nontreated plasma (35, 40, and 18% loss, respectively). The quantity and quality of VWF as well as VWF multimer patterns were not affected by the inactivation method. CONCLUSION: Cryoprecipitates prepared from amotosalen‐treated FFP contained significantly reduced levels of fibrinogen, FVIII, and ADAMTS‐13. However, the VWF quantity and quality was well preserved.     

Comparison and stability of ADAMTS13 activity in therapeutic plasma products

BACKGROUND: The von Willebrand factor (VWF)-cleaving protease, ADAMTS13, is often deficient in cases of thrombotic thrombocytopenic purpura (TTP). The primary treatment of TTP is therapeutic plasma exchange (TPE) utilizing a variety of plasma products that help restore ADAMTS13 activity. However, multiple replacement products are available to choose from. Thawed plasma products have a variable refrigerated shelf life depending on the product type; stability of ADAMTS13 in thawed products stored at 1 to 6 degrees C has not been determined. STUDY DESIGN AND METHODS: ADAMTS13 activity was measured in three types of plasma products and cryoprecipitate. Fresh-frozen plasma (FFP) aliquots and cryoprecipitate-poor plasma (CPP) products were produced from 10 whole-blood (WB) donations. Twenty-four-hour plasma products were manufactured from 10 additional WB donations. ADAMTS13 activity in these products at time of thaw and after 5 days of storage at 1 to 6 degrees C was measured with a modified version of the FRETS-VWF73 fluorogenic assay. ADAMTS13 activity at time of thaw was measured in 10 units of cryoprecipitate and five related CPP products. RESULTS: ADAMTS13 is present in similar amounts in FFP, CPP, and 24-hour plasma products. Storage at 1 to 6 degrees C for up to 5 days did not significantly diminish ADAMTS13 activity. The concentration of ADAMTS13 in cryoprecipitate was significantly higher than that observed in plasma products. CONCLUSION: FFP, CPP, and 24-hour plasma products should be equally effective for ADAMTS13 restoration through TPE and should remain so for the duration of the shelf life of the thawed products.     

The quality of fresh-frozen plasma produced from whole blood stored at 4 degrees C overnight

BACKGROUND: The aim of this study was to assess whether the quality of FFP produced from whole blood stored at 4 degrees C overnight is adequate for its intended purpose. STUDY DESIGN AND METHODS: Fresh-frozen plasma (FFP) separated from whole blood (n = 60) leukodepleted (LD) after storage at 4 degrees C overnight (18-24 hr from donation, Day 1 FFP) was compared with that LD within 8 hours of donation (Day 0 FFP, the current standard method). RESULTS: In more than 95 percent of Day 1 FFP units, levels of factor (F) II, FV, FVII, FVIII, F IX, FX, FXI, and FXII were greater than 0.50 U per mL except for von Willebrand factor (VWF) antigen and FVIII, where 92 and 87 percent of units, respectively, contained greater than 0.50 IU per mL. Compared with historical data on FFP stored for 8 hours, fibrinogen, FV, FVIII, and FXI were reduced by 12, 15, 23, and 7 percent, respectively, but other factors were not significantly reduced. Levels of VWF-cleaving protease activity were not different between FFP prepared from paired units of blood (n = 3) held for 8 or 24 hours, but were below the reference range in an additional 2 of 6 units held for 24 hours. The activities of protein S, protein C, antithrombin III, and alpha(2)-antiplasmin were reduced by less than 10 percent in Day 1 FFP (n = 20), but with final levels above the lower limit of the normal range in greater than 95 percent of units. Activated FXII antigen was not significantly raised in plasma stored for 18 to 24 hours, but levels of prothrombin fragment 1 + 2 were slightly increased (0.88 ng/mL, 18-24 hr; 0.65 ng/mL, < 8 hr). CONCLUSION: These data suggest that there is good retention of relevant coagulation factor activity in plasma produced from whole blood stored at 4 degrees C for 18 to 24 hours and that this would be an acceptable product for most patients requiring FFP.     

The impact of the intensity of serial automated plasmapheresis and the speed of deep-freezing on the quality of plasma

BACKGROUND: Data are lacking on the impact that the intensity of serial donor plasmapheresis has on the quality of source plasma. A study was conducted to examine the quality of source plasma produced by intensive plasmapheresis and slow deep-freezing and to compare it to source plasma manufactured by moderate plasmapheresis and rapid freezing. STUDY DESIGN AND METHODS: Seventy-five plasma samples from intensive plasmapheresis programs (Group 1) and 75 plasma units from moderate plasmapheresis programs (Group 2) were examined. The plasma had been deep-frozen either slowly at -30 degrees C in walk-in freezers (Group 1) or rapidly within 1 hour to a core temperature below -30 degrees C (Group 2). Determinations were made of the plasma levels of citrate; total protein; albumin; IgG; fibrinogen; factors II, V, VII, VIII, and IX; vWF; antithrombin; protein C; D-dimers; and prothrombin fragments 1+ 2. RESULTS: Plasma units of Group 2 contained substantially greater levels of citrate, IgG, FVIII, and FV than samples of Group 1 (p<0.0001). Plasma levels of total protein, albumin, and fibrinogen also were higher in Group 2 (p<0.0001, p = 0.007, and p = 0.006, respectively). Neither plasmapheresis intensity nor freezing procedure had any influence on the levels of factors II, VII, and IX, antithrombin, or protein C. There was no evidence of substantial coagulation activation in the plasma units of either group. However, higher FVIII clotting activity/chromogenic substrate activity ratios in rapidly frozen plasmas and a significant correlation between these ratios and prothrombin fragment 1+ 2 levels suggest that rapid freezing yields both more native FVIII and greater partial activation of FVIII. CONCLUSION: Source plasma collected from donors undergoing intensified plasmapheresis contains markedly lower levels of IgG than plasma units produced by moderate serial plasmapheresis. The combination of intensified plasmapheresis and slower freezing of source plasma results in substantially lower levels of FV and FVIII than does moderate plasmapheresis with rapid freezing. Prospective studies should establish the optimum conditions required for the safe and economic production of source plasma for fractionation.     

Cryoprecipitate prepared from plasma treated with methylene blue plus light: increasing the fibrinogen concentration

When cryoprecipitate is prepared from plasma which has been treated with methylene blue plus light (MB) for the purpose of virus inactivation, clottable fibrinogen content is 40% lower compared with units prepared from untreated plasma. Initial studies showed that when frozen MB plasma units were removed to +2 to +6 degrees C for 4 h and then returned to -40 degrees C prior to cryoprecipitation, fibrinogen recoveries increased from 24 to 42%. Although fibrinogen yield improved when plasma units were stored at +2 to +6 degrees C for varying lengths of time, FVIII levels decreased with increasing time. Conditioning for 8 h was studied in more detail. Groups of two plasma units were mixed together, divided into two equal units, frozen/thawed and treated with MB. One of each pair was stored continually at -40 degrees C, whereas the other was removed to +2 to +6 degrees C for 8 h. Samples were assayed for fibrinogen, FVIII, VWF:Ristocetin cofactor activity (RCo), VWF:Ag and VWF:Collagen binding (CB). The cryoprecipitate fibrinogen content increased to a mean of 207 mg unit(-1). VWF:Ag, VWF:RCo and VWF:CB recoveries also increased. FVIII recovery decreased from 50 to 45% (mean 124 iu unit(-1)). Conditioning has been validated for routine production of cryoprecipitate from imported plasma.     

Coagulation parameters in apheresis and leukodepleted whole-blood plasma during storage

BACKGROUND: Thawing fresh-frozen plasma (FFP) may cause delay in delivery, and one approach to circumvent this is to store plasma at +4 degrees C. Thawed plasma is commonly discarded after a few days of storage, owing to the assumption that coagulation factor activity decreases to clinically unacceptable levels. STUDY DESIGN AND METHODS: Eighteen apheresis plasma (AP) units were collected from blood donors. The collected plasma was divided into two equal parts: one part frozen at -74 degrees C as FFP and one part stored at +4 degrees C as fresh liquid plasma (FLP). Thirty-nine units of whole blood (WB) were collected from blood donors and leukodepleted by inline filtration, followed by plasma separation. Twenty plasma units were frozen at -74 degrees C as FFP and 19 plasma units were stored at +4 degrees C as FLP for 28 days. Plasma aliquots were collected before freezing and immediately after thawing FFP and before and during storage of FLP at Days 14 and 28. Factor (F)V, FVIII, D-dimers, and C1-esterase inhibitor levels were assessed. RESULTS: No significant differences in coagulation factor levels were assessed between FLP prepared from AP and FLP prepared from WB. FV and FVIII levels decreased on average 25 and 50 percent, respectively, at Day 14 of storage. C1-esterase inhibitor and D-dimers levels were not affected. CONCLUSION: Leukodepleted apheresis and WB plasma stored for 14 days retain sufficient levels of FV and FVIII activity for maintenance of normal hemostasis and could therefore be considered useful in selected clinical situations.     

Effect of gamma irradiation with 30 Gy on the coagulation system in leukoreduced fresh-frozen plasma

BACKGROUND: The aim of this study was to investigate the effect of gamma irradiation with 30 Gy on the coagulation system in leukoreduced fresh-frozen plasma (FFP). STUDY DESIGN AND METHODS: In 74 FFP units that had been stored for 352 +/- 103 days below -30 degrees C, the following variables were determined in parallel in an irradiated and not irradiated half: prothrombin time (PT); activated partial thromboplastin time (APTT); thrombin time; antithrombin III; protein C; protein S; von Willebrand factor antigen; ristocetin cofactor; plasminogen-alpha(2)-antiplasmin; the coagulation factors fibrinogen, factor (F)II, FV, FVII, VIII, F IX, FX, FXI, FXII, FXIII, and activated factor XII (FXIIa); D-dimer; fibrin monomer; thrombin-antithrombin complex; prothrombin fragment 1 + 2 (F1+2); plasmin-alpha(2)-antiplasmin complexes (PAPs); and platelet factor 4. The FVII activity ratio was assayed to quantify activation of FVII. RESULTS: Irradiation with 30 Gy resulted in a reduction of APTT (35.0 +/- 4.1 sec vs. 34.4 +/- 4.1 sec; p = 0.00000006) and PT (89.8 +/- 8.2% vs. 90.7 +/- 8.0%; p = 0.002) and a significant increase of the activities of the coagulation factors FII, FV, FVII, F IX, FX, and FXII. FVIII activity decreased from 118 +/- 31 to 116 +/- 32 percent (p = 0.02). Activation of the coagulation system was shown by an increase in the FVII activity ratio (1.19 +/- 0.29 vs. 1.31 +/- 0.34; p = 0.0000001), FXIIa (0.81 +/- 0.50 ng/mL vs. 0.90 +/- 0.51 ng/mL; p = 0.006), and F1+2 (1.19 +/- 0.20 nmol/L vs. 1.24 +/- 0.20 nmol/L; p = 0.000005) after irradiation with 30 Gy, whereas an increase of PAP (16.2 +/- 11.5 ng/mL vs. 20.2 +/- 12.0 ng/mL; p = 0.0004) demonstrated activation of the fibrinolytic system. No negative influence of irradiation with 30 Gy on inhibitors of coagulation was observed. CONCLUSION: Gamma irradiation of leukoreduced FFPs with 30 Gy results in a significant but very weak activation of the coagulation and fibrinolytic system in FFPs.     

Changes in coagulation factor activity and content of di(2-ethylhexyl)phthalate in frozen plasma units during refrigerated storage for up to five days after thawing

BACKGROUND: Thawed plasma is typically transfused to supply coagulation factors but factor activity declines during refrigerated storage. Refrigerating thawed plasma for longer than 24 hours could reduce plasma wastage and make plasma more readily available for emergency transfusions. We measured coagulation factor activity and di(2‐ethylhexyl)phthalate (DEHP) concentration in frozen plasma (FP) thawed and stored at 1 to 6°C for up to 5 days. STUDY DESIGN AND METHODS: FP units prepared using “top‐and‐bottom” collection sets were thawed, refrigerated, and sampled aseptically at 0, 24, 72, and 120 hours after thawing (n = 54). Clotting factor activities and prothrombin times (PTs) were measured using an automated coagulation factor analyzer. DEHP was measured by high‐performance liquid chromatography after hexane extraction (n = 11). Unit sterility was confirmed using an automated microbial detection system. RESULTS: Factor (F)V and FVIII, but not FVII, declined significantly within 24 hours. By Day 5, mean losses were 20, 14, and 41%, in FV, FVII, and FVIII, respectively; fibrinogen activity did not change. PT values were prolonged by 9% on Day 5. Mean DEHP levels increased from 22 ppm at thaw to 66 ppm on Day 5. CONCLUSIONS: The bulk of coagulation factor activity losses during storage occurred in the first 24 hours. Coagulation factor activities remaining in FP after 5 days did not differ from those previously reported in similar products frozen within 24 hours of phlebotomy. While DEHP levels in 5‐day‐thawed FP are not of concern for adult patients, for infants, DEHP levels can be minimized by using FP refrigerated for no more than 24 hours.     

失血性休克大鼠凝血因子的变化研究

本研究观察SD大鼠失血性休克过程中凝血因子的变化 ,探讨凝血连锁反应在休克发生、发展过程中的作用机制。制作大鼠失血性休克模型后 ,4 0只大鼠随机分为休克前组、休克 1、2、4、6、8、12和 2 4小时组。分别检测各组假血管性血友病因子 (vWF) ,凝血因子Ⅷ (FⅧ ) ,组织因子 (TF) ,D 二聚体 (D D) ,纤维蛋白原 (FIB)的血浆浓度及观察凝血酶原时间 (PT)、活化部分凝血酶原时间 (APTT)变化。结果表明 :与休克前比较 ,各休克组PT、APTT逐渐延长 ,于休克后 4 - 6小时显著延长 (P <0 .0 0 1) ,APTT平均 5 9.7秒 ,PT平均 30 .2秒。休克后 ,血浆D 二聚体显著增加 (P <0 .0 0 1) ,于休克 8小时达到峰值。血浆vWF、FⅧa、TF、FIB于休克早期增加 ,随着休克的发展 ,纤维蛋白原于休克 2小时开始明显下降 (P <0 .0 0 1) ,休克 12小时降至最低值。TF、vWF、FⅧa分别于休克 6小时、8小时开始明显下降 (P <0 .0 0 1或P <0 .0 5 )。凝血因子消耗比率 :FⅧ为 (86 .1± 1.8% ) ,纤维蛋白原为 (89.6± 0 .6 % ) ,假血管性血友病因子为 (5 5± 1.4 % ) ,组织因子为 (6 2± 2 .5 % ) ,其中 ,FⅧ、纤维蛋白原消耗比例较大。失血性休克时以外源性凝血途径激活为主 ,内源性凝血途径作用较小。纤维蛋白原和D 二聚体、PT、APTT可用     

Rapid controlled thawing of fresh-frozen plasma in a modified microwave oven

A microwave oven has been specifically modified to permit rapid thawing of fresh-frozen plasma (FFP) by using a rotating disc with a temperature sensor to hold the plasma bag. This modification makes it possible to mix the FFP continuously during thawing, and automatically shuts the oven off when the plasma reaches 21 degrees C. Comparisons were made between FFP thawed in the modified microwave oven and FFP thawed conventionally in a 37 degrees C waterbath. The following tests were done: total protein, albumin, and immunoglobulin concentrations; plasma fibrinogen, factor VIII, and factor IX activities; protein electrophoresis, albumin aggregation, hemolytic complement activity, and plasma particle count and size. In no case was there a significant difference between plasma thawed in the microwave oven compared with that thawed in the waterbath. Further, microwave thawing was reliable and rapid; all units of FFP thawed in less than 6 minutes, and the thawed plasma did not vary by more than 6 degrees C from the preselected final temperature of 21 degrees C. Thus, it appears that controlled thawing of FFP in a microwave oven specifically designed for this purpose is an effective and reliable method and has many advantages over conventional thawing of FFP.     

Refreezing previously thawed fresh-frozen plasma. Stability of coagulation factors V and VIII:C

With the growth in autologous blood programs and the increased scrutiny of the indications for transfusion of fresh-frozen plasma (FFP), an increase has been seen in the number of occasions on which FFP was requested and thawed but then not transfused. The coagulation properties of FFP units that were refrozen and then rethawed were therefore studied. Fifty-eight units of plasma were studied, with each experimental unit of FFP paired with an identical control unit. Experimental units were frozen, stored at -65 degrees C, thawed, stored at 1 to 6 degrees C for various periods of time up to 24 hours, and then refrozen, stored at -65 degrees C, rethawed, and stored again in the refrigerator for up to 24 hours. Control units were frozen once at the time the experimental units were first frozen and thawed once at the time of the second thaw of the experimental units. Aliquots of plasma were sampled periodically and were later batch-tested for prothrombin time (PT), activated partial thromboplastin time (aPTT), and factor V and VIII:C activity. The results of coagulation testing of the twice-frozen plasmas were always within the normal range. There was a slight but statistically valid prolongation of the PT and aPTT and a decrease in the factor V and VIII:C levels for twice-frozen plasma compared with control plasma. The greatest decline occurred in the level of factor VIII:C. The measured deterioration in coagulation of twice-frozen FFP is unlikely to be of clinical importance. Refreezing FFP may eventually prove useful for rare donor, autologous, and massive transfusion programs.     

新的双重杂合性FV基因突变导致的重型遗传性FV缺陷症

目的对一个遗传性凝血因子V(FV)缺陷症家系进行FV基因突变的检测。方法用活化部分凝血活酶时间(Am)，凝血酶原时间(PT)及FV促凝活性(FV：C)和FV抗原(FV：Ag)测定进行表型诊断；用PCR法对先证者FV基因25个外显子及其侧翼序列进行扩增，PCR产物纯化后直接测序，检测其基因突变。突变位点经限制性内切酶分析证实：结果先证者APTT 249．2s．PT46．6s。TT17．9s，Fg3．42g/L，FV：C0．1％，FV：Ag 1．5％，FⅡ：C99％，FⅦ：C110％、FⅧ：C95％、FⅨ：C88％、FX：C120％、vWF121％；FV外显子区共发现4个与基因文库Z99572序列不同的位点，其中位于exon13区的杂合性2238～2239de1AG导致移码突变和终止密码子的提前m现(Asp689stop)，位于exon23区的杂合性G6410T错义突变导致Gly2079 Val.家系分析表明前者遗传于母亲，后者遗传于父亲。结论2238～2239delAG导致的移码突变和G6410T导致的错义突变．是导致先证者FV缺陷的原因。这是2个导致遗传性FV缺陷症的新的FV基冈突变位点。     

Conversion to the buffy coat method and quality of frozen plasma derived from whole blood donations in Canada

BACKGROUND: Canada converted from the platelet‐rich plasma (PRP) method to the buffy coat (BC) method of processing whole blood donations between 2006 and 2008. We measured coagulation variables in plasma units during this transition, in 2006 (PRP only), 2007 (BC and PRP), and 2008 (BC only) to test the hypothesis that this conversion would not affect frozen plasma (FP) quality. STUDY DESIGN AND METHODS: Fresh‐frozen plasma (FFP; frozen within 8 hr of collection) or FP (frozen within 24 hr of collection) units were shipped on dry ice from 12 plasma manufacturing sites, thawed, and characterized using an automated coagulation analyzer, at a single testing site. RESULTS: FP made by the BC method (FP‐BC) exhibited fibrinogen, Factor (F)V, ABO‐matched FVIII, and antithrombin levels at least as high as FP made by the PRP method (FP‐PRP) and supported global clotting, as measured by prothrombin time or activated partial thromboplastin time, to an indistinguishable extent as FP‐PRP. FP‐BC and FP‐PRP did not differ in ABO‐matched FVIII levels, but both contained 30% to 35% less FVIII than FFP. There was no discernible effect of the site of manufacturing on plasma quality. FP‐BC units leukoreduced by centrifugation contained more FV activity than those leukoreduced by filtration, but the difference was unlikely to be of clinical significance. CONCLUSION: Our data suggest that no reduction in FP quality, at least in the characteristics we tested, accompanied the switch from the PRP to the BC method processing of whole blood donations in Canada.