Anticoagulant Citrate Phosphate Double Dextrose Solution

Studies of the effectiveness of 250mL CP2D and 250mL AS-3 made by Haemonetics
Corporation Union, South Carolina facility included in vivo survival data in healthy
subjects at one test site and in vitro characterization of stored red blood cells at
three test sites.

Haemonetics Clinical Evaluation #95012, involved autologous in vivo recovery and
survival studies, and in vitro red blood cell characterization conducted at a single
site. RBC results collected with Haemonetics CP2D/AS-3 were compared to crossover
controls of manually collected RBCs from the same donors using CP2D/AS-3 made by
Medsep Corporation (crossover manual controls). In addition, results were
compared to data from other RBC apheresis donors using CP2D/AS-3 solutions
manufactured by MedSep (unmatched apheresis controls). Test parameters included
hematocrit, hemoglobin, RBC and WBC counts, ATP levels, pH, supernatant
potassium, free hemoglobin, supernatant glucose levels, and product weight. The
RBC quality of test units after 42-day storage was equivalent or slightly superior to
those of the control units. RBCs collected by the two different collection protocols
(Red Blood Cells with or without Plasma) showed no significant differences in terms
of RBC quality and red blood cells from all groups met FDA and AABB guidelines for
24-hour recovering at 42 days storage.

Study #98001 was performed using 250mL CP2D and 250mL AS-3 to confirm, in
vitro, the results of Clinical Evaluation #95012. Three test sites each performed three
RBCP and three 2RBC apheresis procedures using the MCS+ LN8150 and associated
collection sets. In addition, each site collected six manual whole blood units as
concurrent (unmatched) controls using Medsep sets and prepared RBCs according to
standard procedures.

In vitro tests of red blood cell functional and physical integrity were conducted at
Days 0 and 42 of storage. All plasma units from the RBCP and manual procedures
were evaluated on Day 0 for in vitro product characteristics. Group t-tests (two
tailed) were performed to examine any statistically significant differences between
the properties of apheresis and manual RBCs. Statistically significant differences
(p<n; 0.05) were evaluated in terms of clinical significance. Differences in glucose or
lactate levels, hematocrit, pH, and supernatant potassium were not considered
clinically relevant because these parameters have not been found to correlate
significantly with the in vivo 24 hour percent recovery.

The results from this study confirmed the observations from Clinical Evaluation
#95012. The quality of stored RBC products collected by apheresis is similar to RBC
products collected manually. A difference in ATP and hemolysis levels, both after
processing and on day 42 of storage, for the apheresis units were not statistically
significant when compared to ATP and hemolysis levels from manually collected
RBCs and were similar to historicai reference values.

AS-3 300ml was used in conjunction with an automated closed system, the
Haemonetics Model 215 (now called the ACP) in Clinical Evaluation #97002. Five test
sites were used in this study. 100 samples were tested in vitro and 30 in vivo. Red
blood cells derived from CPDA-1 whole blood units were used. AS-1 red blood cells
were used as a control. A total of 140 red blood cell units were glycerolized and
deglycerolized. The in vitro RBC quality and in vivo RBC viability data obtained on
these units demonstrate that red cell units glycerolized and deglycerolized using the
Model 215 System and resuspended in 300ml AS-3 solution manufactured by
Haemonetics, Union, South Carolina are processed in a closed system and can be
stored for 15 days at 4°C.

Clinical report TR-CLN-100229-A was conducted to support the use of CP2D for the
collection of FFP and PF24 {plasma collected and stored Plasma collected using the
822, 822-2P and 822F-2P disposable sets may be frozen within 8 hours (FFP) or
within 24 hours which includes 8 hours room temperature storage and 16 hours
refrigeration storage (PF24). The following tables provide a summary of the results
of this evaluation.

Note: Table 1, Table 2, and Table 3 below demonstrate the results of a clinical
study conducted for various coagulation factors on plasma that was frozen within
8 hours (FPP) versus plasma frozen within 24 hours, which includes 8 hours
room temperature storage and 16 hours refrigeration storage (PF24).

As noted in Table 1 below, Significant differences were observed between PF24 and
FFP for assays Prothrombine Time, Partial Thromboplastin Time, FVlla, and
FVIII.

Table 2 provides the results for Plasma Sample A, which was an outlier and
subsequently excluded from the analysis.

Table 3 provides a summary of how many samples tested in each group differed by
>20% when FFP and PF24 were compared.

Table 1, Summary of FFP and PFZ4 plasma product assays (N=59)

Assay	Mean (SD)		Median		(Min, Max)		Mean difference (PF24, FFP) (95% confidence interval)
	FFP	PF24	FFP	PF24	FFP	PF24
Prothrombin Time (sec)	12.0 (0.50)	12.1 (0.59)	11.9	12.1	(11.2, 13.8)	(10.5, 14.0)	015 (0.09,0.20)
Activated Partial Thromboplastin Time (sec)	29.4 (3.32)	30.3 (3.34)	28.8	29.8	(23.1, 38.5)	(24.1, 39.5)	0.86 (0.64, 1.08)
Favtor V (%)	120.1 (23.93)	120.0 (23.88)	120.0	120.0	(61.0, 181.0)	(59.0, 182.0)	-0.14 (-2.03, 1.76)
Factor Vlla (mU/mL)*	60.5 (26.95)	45.0 (24.72)	54.0	41.5	(21.0, 131.0)	(12.0, 159.0)	-15 .50 (-19 .23, -11.77)
Factor VIII:C (%)*	94.3 (34.24)	81.7 (27.31)	88.5	82.5	(33.0, 194.0)	(34.0, 152.0)	-12.67 (-15.97, -9.38)
AT-III (%)	97.8 (12.24)	94.7 (15.97)	98.0	95.0	(71.0, 126.0)	(19.0, 134.0)	-3.05 (-6.30, 0.20)
Factor XI (%)	112.2 (17.42)	112.8 (18.46)	113.0	113.0	(81.0,151.0)	(82.0, 155.0)	0.63 (-0.70,1.95)
vWF (R:Co) (% function)	89.5 (35.65)	89.4 (35.58)	85.0	87.0	(35.0, 179.0)	(37.0, 185.0)	-0.15 (-2.21, 1.91)
Protein S (% activity)	74.9 (19.61)	73.4 (20.76)	72.0	71.0	(33.0, 125.0)	(39.0, 126.0)	-1.49 (-3.81, 0.83)
Protein C (% activity)	117.1 (17.59)	117.7 (17.49)	115.0	118.0	(65.0,157.0)	(68.0, 162.0)	0.53 (-0.54,1.59)
Fibrinopeptide F 1.2 (pmol/L)*	136.4 (63.34)	133.1 (55.15)	113.0	114.0	(80.0, 344.0)	(80.0,331.0)	-3.29 (-8.37,1.78)
Thrombln-Anti- thrombin Complex (ng/mL)	2.3 (1.38)	2.1 (0.40)	2.0	2.0	(2.0, 12.4)	(2.0,4.8)	-0.17 (-0 .53, 0.18)

*Sample size = 58. The outlier pair was excluded from the analysis based on the statistical criteria or laboratory errors.

One sample (Plasma Sample A) showed significantly increased levels for Factor VIIa and F1.2. Another sample (Plasma Sample B) showed significantly decreased level of Factor VIII:C for the FFP sample (Table 2). These data were excluded from Table 1 since the values for VIIa and F1.2 were more than 4 SD from the mean. The factor VIII:C was not more than 4 SD from the mean but was significantly lower in FFP as compared to PF24. While other parameters in this sample did not show significant differences when compared to the control FFP, the FDA could not exclude the possibility that the activation found in the PF24 sample was related to the preparation procedure. The rate of such event occurrence was unknown and, based on review of published values for PF24 products, is most likely very rare. The clinical significance of the elevated markers of clotting activation for transfusion recipients is undetermined.

Table 2, Three outlier pairs in plasma measurements

Subject #	Measurement	FFP	PF24	Range	without	the	outliers
				FFP Min	FFP Max	PF24 Min	PF24 Max
RBCP16	Factor FVIIa (mU/mL)	220	767	21	131	12	159
RBCP16	F1.2 (pmol/L)	702	1940	80	344	80	331
RBCP01	Factor VIII: C(%)	2	63	33	194	34	152

Table 3, Proportion of Paired Sample:PF24 is greater than or less than FFP by more than 20%

Assay	PFZ4>FFP by more than 20%	PF24<FFP by more than 20%
Prothrombine Time (sec)	0% (0/59)	0% (0/59)
Partial Thromboplastin Time (sed	0% (0/59)	0% (0/59)
Factor V (%)	1.7% (1/59)	0% (0/59)
Factor Vila (mU/mL)	3.4% (2/58)	89.7% (52/58)
AT-III (%)	1.7% (1/59)	8.5% (5/59)
Factor VIII :C (%)	1.7% (1/58)	22.4% (13/58)
Factor XI (%)	0% (0/59)	0% (0/59)
Fibrinopeptide 1.2 (pmol/L)	0%(0/58)	1.7% (1/58)
Protein C (% activity)	0% (0/59)	0% (0/59)
Protein S (% activity)	5.1% (3/59)	1.7% (1/59)
Thrombin-Antithrombin Complex (ng/mL)	1.7% (1/59)	3.4% (2/59)
vWF (% activity)	3.4% (2/59)	1.7% (1/59)

Note: Plasma collected may be labeled as FFP or Plasma Frozen within 24 Hours (PF24).

Note: For FFP, the plasma should be placed in a freezer within 8 hours from the completion of collection. The plasma should be frozen at -18°C or colder.
For product intended to be labeled as PF24, the plasma should be refrigerated within 8 hours of collection at 1 – 6°C and frozen within 24 hours of collection.
FFP and PF24 should be frozen at -18°C or colder.