Date:  March 14, 1996 
From:    Director, Center for Biologics Evaluation and 
Research 
Subject:  Additional Recommendations for Donor Screening With 
a Licensed Test for HIV-1 Antigen 
To:    All Registered Blood and Plasma Establishments 
INTRODUCTION
 
On August 8, 1995, FDA issued a memorandum to all registered
 
blood and plasma establishments entitled, "Recommendations for
 
Donor Screening with a Licensed Test for HIV-1 Antigen".  This
 
memorandum supplements that memorandum and provides additional
 
recommendations regarding storage of samples for HIV-1 antigen
 
testing as well as clarifications of the previous
 
recommendations regarding specific implementation issues. 

 
STORAGE OF SAMPLES FOR HIV-1 ANTIGEN TESTING
 
Concerns have been raised about the stability of HIV-1 antigen 

containing samples during shipping and storage.  Stability
 
data from studies of both antibody(-) and antibody(+)
 
specimens from seroconversion series which were performed by
 
manufacturers in support of product license applications for
 
HIV-1 p24 antigen test kits suggest that detectable antigen
 
levels in some serum or plasma specimens may be adversely
 
affected by elevated temperatures and long periods of storage. 

The available data have not demonstrated false-negative
 
results in the HIV-1 antibody test if antigen(+)/antibody(+)
 
samples are stored under conditions that would allow
 
significant antigen-antibody complexing or antigen
 
deterioration to occur.  The following is a summary of the
 
findings on antigen stability:
 
   
All studies confirmed the stability of HIV-1 p24 antigen(s) in
 
antigen(+)/antibody(-) samples.  Only one sample showed a loss
 
of reactivity following storage for three days at 4EC and at
 
26EC.  When this sample was stored for longer periods, no
 
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  further decrease in reactivity was observed and the sample
 
remained positive for HIV-1 p24 antigen(s). 

Storage of some antigen(+)/antibody(+) samples for as little
 
as three days at 4EC and at 26EC resulted in a loss of
 
reactivity on the antigen screening test, including the
 
occurrence of false negatives.  When testing was not completed
 
within three days of sample collection with storage at 4EC, an
 
increased likelihood of failure of initially reactive samples
 
to be repeatedly reactive on the antigen screening test,
 
invalid neutralization test results, and false negative
 
neutralization test results occurred. 

Instability of the antigen signal in both
 
antigen(+)/antibody(-) and antigen(+)/antibody(+) samples held
 
at 37EC has been clearly demonstrated, suggesting that
 
antigen-antibody complexing or antigen deterioration or both
 
occur to a greater extent at such elevated temperatures. 

Overall, the preponderance of the available data suggests that
 
most antigen(+) samples, especially antigen(+)/antibody(-)
 
seroconversion samples which are of critical importance for
 
the HIV-1 p24 antigen test to detect in donor screening, are
 
stable at 4EC for at least seven days and that exposure to
 
room temperatures below 26EC can be tolerated for up to three
 
days.  These findings are the basis for the attached
 
recommendations on sample storage prior to antigen testing.
 
CLARIFICATIONS OF PREVIOUS RECOMMENDATIONS
 
1.  FDA has been asked to clarify its expectations related to
 
the "date of implementation" of the test for HIV-1 antigen(s).
 
In Recommendation A.1. of the August 8, 1995, memorandum, the
 
FDA stated that, "all donations of Whole Blood, blood
 
components, Source Leukocytes and Source Plasma should be
 
screened for HIV-1 antigen(s) by an FDA licensed test labeled
 
specifically for use in donor screening," and that, "...this
 
recommendation should be implemented within three months of
 
the commercial availability of the first such test.  Following
 
the date of implementation, only units from donors found
 
negative on HIV-1 antigen screening ... should be released..."
 
CLARIFICATION:  The "date of implementation" is the date
 
within three months of HIV-1 antigen test licensure that a
 
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  manufacturer declares to FDA will be the date following which
 
all units released by the establishment will have been
 
screened for HIV-1 antigen(s).
 
2.  FDA has been asked to clarify how to deal with Source
 
Plasma and Recovered Plasma in inventory for further
 
manufacturing.  Recommendation A.2. states, "Within three
 
months of the commercial availability of the first FDA
 
licensed HIV-1 antigen test labeled specifically for use in
 
donor screening, all inventoried units ... available for
 
release should be screened for HIV-1 antigen(s)." 
   
CLARIFICATION: For Source Plasma and recovered plasma, this
 
recommendation is intended to apply only prospectively, i.e.,
 
to units drawn after the implementation date (within three
 
months of licensure of the first donor screening test for HIV
1 antigen).  Units collected on or after the implementation
 
date should be tested for HIV-1 antigen and labeled as tested
 
and found negative.  Untested units collected prior to the
 
implementation date may be distributed without special
 
labeling.  Additional labeling consistent with section B.3. of
 
the August 8, 1995 memorandum may be used at the
 
establishment's discretion.
 
3.  FDA has been asked whether the three month time frame for
 
implementation of the antigen test also applies to inventories
 
held by consignees.  Recommendation A.3. states that, "Blood
 
establishments should cooperate with consignees to insure that
 
inventoried within-date units intended for use in transfusion
 
which were distributed prior to test implementation are either
 
replaced with screened units or else tested for HIV-1
 
antigen(s) as soon as feasible."
 
CLARIFICATION:  These units are in hospital inventories and
 
otherwise out of the control of the blood establishment. 

Blood establishments should work with transfusion services and
 
other consignees to complete inventory testing or exchange to
 
the extent possible and as soon as it is feasible.  FDA
 
recognizes that this may not occur within the three month
 
period recommended for test implementation. This
 
recommendation does not apply to units already released for
 
transfusion.
 
4.  FDA has been asked to explain the rationale for imposing
 
controls on the distribution of autologous units for
 
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  transfusion in the event of a repeatedly reactive screening
 
test for HIV-1 antigen in the patient/donor.
 
CLARIFICATION:  The procedures described in section B.2.a.
 
pertaining to autologous units should be applied to any unit
 
intended for autologous use that tests repeatedly reactive on
 
the HIV-1 antigen screening test.  This recommendation has
 
been made because of the possibility of false negative
 
neutralization tests.
 
5.  FDA has been asked to explain its position on the
 
regulatory implication of any follow-up testing performed on a
 
donor prior to eight weeks after a repeatedly reactive
 
screening test for HIV-1 antigen, and the Agency expectation
 
regarding deferral and reentry in the case that a donor has a
 
repeatedly reactive screening test for HIV-1 antigen on more
 
than one independent collection.  Recommendation C.2. states
 
that, "Donors whose blood samples are INDETERMINATE based on
 
the neutralization test or 

untested by neutralization should be temporarily deferred from
 
donation for a minimum of eight weeks."   Recommendation C.2.a.
 
further states that "The donor can automatically be
 
reinstated...if all donor screening tests (including HIV
 
antigen) are performed and found NEGATIVE, and the donor meets
 
all other suitability criteria." 
 
CLARIFICATION:  Blood or plasma centers wishing to perform
 
follow-up testing on donors who are temporarily deferred may
 
do so prior to the end of this eight week period for donor
 
notification purposes or for medical reasons. However, if the
 
donor is retested prior to the end of the eight week deferral
 
period and repeatedly reactive EIA results are again obtained,
 
the donor should be permanently deferred.  A negative result
 
on the EIA or neutralization test may be used in donor
 
counseling, however, only a negative screening test result
 
obtained at least eight weeks after the latest repeatedly
 
reactive test qualifies as the test of record for purposes
 
of donor reentry.
 
6.  FDA has also been asked to clarify recommendation C.2.b.
 
(the donor is permanently deferred if the screening test for
 
HIV antigen is repeatedly reactive on any subsequent
 
evaluation or donation...) when a reinstated donor tests
 
repeatedly reactive on a donation subsequent to having been
 
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  reinstated. 

CLARIFICATION: The donor should be temporarily deferred again
 
and is eligible for reentry eight weeks later as outlined in
 
C.2.a.
 
The recommendations contained in this memorandum may be
 
implemented without prior approval from FDA.  However,
 
licensed establishments implementing these recommendations
 
should submit by official correspondence a statement to their
 
product license file indicating the date that revised standard
 
operating procedures consistent with these recommendations
 
have been established and implemented.  If an establishment
 
believes that an alternative approach would provide equivalent
 
protection, the establishment is invited to discuss the
 
approach with FDA for FDA's evaluation.
 
The recommendations for storage and testing discussed in this
 
memo apply only to HIV-1 antigen testing.  In the event that
 
an FDA approved screening test may require different storage
 
conditions, the package insert instructions should be
 
followed.
 
Although this guidance document does not create or confer any
 
rights, privileges, or benefits for or on any person and does
 
not operate to bind FDA or the public, it does represent the
 
agencys current thinking with regard to HIV-1 antigen
 
screening.
 
Questions and comments about the recommendations or
 
clarifications contained in this memorandum may be directed to
 
the Division of Transfusion Transmitted Diseases, FDA/CBER
 
(HFM-310), 1401 Rockville Pike, Rockville, MD 20852; FAX: 

(301) 480-7928.
 
_______________________ 
Kathryn C. Zoon, Ph.D.
 
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  Additional Recommendations for Donor Screening
 
With a Licensed Test for HIV-1 Antigen
 
March 14, 1996
 
The following recommendations regarding storage of donor
 
samples prior to testing for HIV-1 antigen(s) by licensed
 
tests are intended to supplement those contained in FDA's
 
memorandum to all registered blood and plasma establishments
 
dated August 8, 1995.  In order to assure the integrity of
 
donor samples to be tested for HIV-1 antigen(s), the FDA
 
recommends the following: 

1.  All testing for HIV-1 antigen(s), including the initial
 
screening test, the repeat test, and the neutralization test
 
for repeatedly reactive specimens, should be completed as soon
 
as feasible following sample collection.
 
2.  Serum or plasma samples for testing for HIV-1 antigen(s)
 
should be stored for no longer than seven days at a
 
combination of room temperature (15-26o C
 ) and nominal 4EC
 
(generally accepted as 2-8 oC
  for laboratory samples) following

sample collection.  This seven day period of storage without
 
freezing should include no more than three days at room
 
temperature.
 
3.  Samples that have to be stored for more than seven days
 
prior to testing should be kept frozen at -20o C
  or lower.  No
 
more than five freeze thaw cycles should be performed on any
 
sample prior to testing for HIV-1 antigen.
 
4.  Samples should not be exposed to temperatures above 26EC
 
during shipping, storage or thawing from the frozen state. 

FDA encourages blood and plasma establishments to ship samples
 
by methods shown to maintain temperatures at 2-8EC.  For
 
shipping at ambient temperatures, shipping conditions should
 
assure that temperatures do not exceed 26EC.
 
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