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US Public Health Service Guidelines for Testing and Counseling Blood and Plasma Donors for HIV-1 Antigen

US Public Health Service Guidelines for Testing and Counseling Blood and Plasma Donors for HIV-1 Antigen

In the United States, the implementation of antibody testing in
1985 of all donated blood for human immunodeficiency virus type
1 (HIV-1) resulted in a substantial decrease in the transmission
of HIV through blood transfusions. To further decrease the risk
for transmission of HIV by transfusion, the testing of all blood
donations with a combination antibody test for HIV-1 and HIV type-2
(HIV-2) was implemented in June 1992.

The risk for HIV transmission by transfusion of screened blood
is minimal. Nearly all cases of transfusion-associated HIV transmission
are now caused by blood donated during the infectious window period
(ie, when recently infected donors are infectious but have not
yet developed detectable levels of HIV antibody). When whole-virus-lysate
enzyme immunosorbent assays (EIAs) were used to screen blood donations
from 1985 through 1990, the average length of the window period
was 45 days (95% confidence interval [CI] = 34 to 55 days). The
average window period of the most sensitive contemporary recombinant
protein-based EIA for HIV-1 and HIV-2 antibodies is now 20 days
less, yielding an average infectious window period of 25 days
(95% CI = 9 to 41 days).

The increased sensitivity of contemporary HIV-antibody EIAs, improved
donor interviewing about behaviors associated with risk for HIV
infection, and deferral of donors who test positive for HIV, hepatitis,
human T-cell leukemia virus type 1 (HTLV-I), or syphilis have
considerably improved the safety of the US blood supply. In 1993,
only approximately 6 per 100,000 blood donations collected by
the American Red Cross tested positive for HIV antibody. In addition,
only an estimated 1 in 450,000 to 1 in 660,000 donations per year
(ie, 18 to 27 donations) were infectious for HIV but were not
detected by current screening tests.

During the acute period of infection, tests for p24 antigen can
detect HIV infection earlier than antibody tests. P24 antigen,
the core structural protein of HIV, is detectable 2 to 3 weeks
after HIV infection during the initial burst of virus replication
associated with high levels of viremia. During this time, the
blood of infected persons is highly infectious, and tests for
p24 antigen are usually positive. On average, p24 antigen is detected
an estimated 6 days before antibody tests become positive. When
antibodies to HIV become detectable, p24 antigen is often no longer
detectable because of antigen-antibody complexing and viral clearance.

In August 1995, the FDA recommended that all blood and plasma
donations be screened for p24 antigen, effective within 3 months
of licensure of a test labeled for such use. The FDA recommended
p24 screening as an additional safety measure because (1) recent
studies indicated that p24 screening reduces the infectious window
period; (2) implementation of p24-antigen testing had become logistically
feasible for mass screening; and (3) such testing would reduce
the risk for HIV infection for persons who receive donated blood
or blood products. Among the 12 million annual blood donations
in the United States, p24-antigen screening is expected to detect
4 to 6 infectious donations that would not be identified by other
screening tests. If each of these units were divided into an average
of 1.8 blood components, antigen testing would result in removal
of an estimated 7 to 11 infectious components each year that would
otherwise be available for transfusion. The FDA regards donor
screening for p24 antigen as an interim measure pending the availability
of technology that would further reduce the risk for HIV transmission
from blood donated during the infectious window period.

This report provides guidelines for (1) interpreting p24-antigen-assay
results, (2) counseling and follow-up of blood and plasma donors
who have positive or indeterminate antigen-test results, and (3)
using p24-antigen testing in settings other than blood banks.
These guidelines may be modified when additional information concerning
antigen testing under mass screening conditions is collected and
analyzed.

p24-Antigen Test Algorithm and Interpretation of Results

The p24-antigen screening assay is an EIA performed on serum or
plasma. If the first screening test is nonreactive, the test result
is reported as negative (see Table 1). If the first screening
test is reactive, the p24 EIA is repeated in duplicate. If both
duplicate tests are nonreactive, the test result is reported as
negative. If at least one of the repeated p24 EIA tests is reactive,
the test is considered repeatedly reactive; the donation is then
discarded, the donor is deferred from donating blood, and a more
specific assay (the neutralization assay) is performed to verify
the presence of p24 antigen. The neutralization assay should be
performed before informing donors of test results.

As specified by the FDA, donations collected within 3 months of
a repeatedly reactive p24-antigen test (regardless of neutralization-assay
results) should be quarantined pending results of repeat donor
testing for antigen and antibody to HIV. Sample storage requirements
and time restrictions specified in the test kit package insert
should be closely followed to prevent sample deterioration, and
thus, invalid test results. The FDA recommends that units of whole
blood, blood components, source leukocytes, and source plasma
obtained from donors whose blood samples are repeatedly reactive
on p24-antigen screening tests be destroyed or quarantined and
not used for transfusion or for manufacturing into injectable
products.

Available data indicate that the p24-antigen assay is sensitive
and specific. The specificity of the p24-antigen test was calculated
by two test-kit manufacturers to be 99% to 95% and 99.93% (Table
2
). In addition, in one study of 514,000 donations, 225 were repeatedly
reactive on the screening test. Of these donations, neutralization
tests were negative for 220 (98%). Five (2%) donations were negative
on neutralization tests and had detectable HIV antibodies. Testing
by the polymerase chain reaction (PCR) for HIV DNA and RNA was
performed on 120 of these nonneutralizing blood donations, all
of which were negative for HIV. Follow-up samples were obtained
from 79 of these donors, all of which were negative for HIV-1
antibody.

In a prospective study conducted from September 1993 through September
1995, a total of 305,989 donations were tested for p24 antigen;
3 donors had both repeatedly reactive p24-antigen EIA screening-test
results and positive neutralization results (2 of whom were also
HIV-antibody positive), and 223 donors had repeatedly reactive
p24-antigen EIA screening-test results and negative neutralization
results. Of those donors who had negative neutralization results,
81 later returned to donate blood again. Of these donors, 65 had
negative test results for HIV-1/HIV-2 antibody and for antigen
EIA and neutralization. However, 16 donors who were HIV-1/HIV-2
antibody negative on subsequent donations continued to have repeatedly
reactive p24-antigen EIA screening tests that did not neutralize.

A recent study of 51 seroconversion panels has yielded an estimate
of the clinical sensitivity of the p24-antigen screening test
in detecting blood donated during the infectious window period.
An analysis of 69 preseroconversion samples that were positive
for HIV-1 RNA by PCR demonstrated that the antigen test was reactive
for 51 (74%) of those samples. To assess the sensitivity of the
neutralization test, two antigen-test-kit manufacturers also performed
neutralization testing on samples of blood from persons seroconverting
to HIV. One manufacturer tested 102 repeatedly reactive specimens
from 30 seroconverting plasma donors; 100% were positive on the
neutralization assay. Similarly, a second manufacturer found that
all 52 repeatedly reactive specimens from 25 seroconverting plasma
donors were positive on the neutralization test.

Donor Counseling, Follow-up, and Deferral

Counseling blood and plasma donors who have positive or indeterminate
HIV-test results is an essential adjunct to HIV testing. Counseling
in the blood-bank setting (1) provides information about follow-up
diagnostic evaluation and available medical, preventive, and psychosocial
services and (2) assists infected persons in preventing transmission
to others. HIV counseling should be conducted in accordance with
PHS standards and guidelines.

PHS guidelines for notification and counseling of donors who have
repeatedly reactive antigen-test results are based on available
data. These guidelines may be modified after the collection and
analysis of additional information concerning antigen testing
under mass screening conditions.

Donors with Positive P24-Antigen Results--Donors whose
HIV-antibody-test results are negative but whose screening-test
results for

HIV antigen are repeatedly reactive and neutralization-assay results
are positive, should be counseled that they are probably infected
with HIV (Figure 1). Donors who have such test results should
be notified promptly after a positive neutralization test. Prompt
notification is important because persons who are newly infected
with HIV and do not have HIV antibodies often have high viral
titers and may be at high risk for transmitting HIV infection.
According to FDA recommendations, donors who have repeatedly reactive
and neutralizing p24-antigen tests should be advised that they
are permanently deferred from future blood and plasma donation.

Donors who have repeatedly reactive EIA and neutralizing HIV antigen
tests should have their results confirmed by follow-up antibody
testing; diagnosis of HIV infection should not be made on the
basis of p24-antigen test results alone. Arrangements for follow-up
antibody testing should be incorporated into routine counseling.
Because the time between detection of antigen and antibody is
estimated to be an average of 6 days, donors who have positive
p24-antigen test results can be offered repeat antigen and antibody
testing at any follow-up visit. If repeat antigen tests remain
reactive and antibody tests remain negative, antibody testing
should be repeated after a minimum of 8 weeks to allow time for
antibodies to develop. Pending repeat testing to confirm the initial
positive antigen test result, strategies to reduce transmission
should be implemented immediately by the donor (eg, abstaining
from sexual intercourse and using condoms consistently and correctly).
If follow-up antibody tests are positive--thus confirming HIV
infection--infected persons should be referred for medical care;
sex and needle-sharing partners of such persons should be advised
to seek HIV testing at clinical sites.

Some donors repeatedly have positive p24-antigen and negative
HIV-antibody-test results, although such an occurrence is unusual.
If after 8 weeks such persons still have negative antibody-test
results, they should be referred for further medical evaluation,
including determining CD4+ T-lymphocyte cell count or percentage.
Testing for HIV by PCR or culture also may be helpful in determining
HIV status; however, neither test is licensed for diagnosis of
HIV infection.

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