Evaluation of Discrepant Infectious Disease Results in Deceased Organ Donors: Insights From a Retrospective Analysis of Post‐Policy Testing

ABSTRACT

Background

In 2021, a new policy was implemented by the Organ Procurement Transplant Network requiring Organ Procurement Organizations to draw a repeat blood sample for deceased organ donors if donation had not proceeded within 96‐h after the initial blood sample for screening was obtained. We performed an analysis of over 2600 deceased donor test results, comparing initial results to repeated blood draw results for human immunodeficiency virus, Hepatitis B virus, and Hepatitis C virus serology and nucleic acid test (NAT) tests. This study reviews result discrepancies and explores investigations behind peculiar results.

Methods

Infectious disease results from deceased organ donors were analyzed retrospectively for this study. Donor specimens were collected throughout the United States and tested at eleven laboratories. Food & Drug Administration‐approved donor screening tests were used to determine donor eligibility.

Results

There was a 1.69% discrepancy rate comparing results from repeat blood draw specimens to original specimen results. Of these discrepancies, 0.75% of the donors had results (enzyme‐linked immunoassay and/or NAT) that changed from non‐reactive to reactive. 0.68% of donors had results that changed from reactive to non‐reactive. 0.26% of results changed from Ultrio repeatedly reactive, non‐discriminated to either non‐reactive or reactive.

Conclusion

This study represents that there is more than a 1% chance that discrepant results will be obtained. Despite the low incidence of discrepancies, these rare occurrences can complicate clinical decision‐making, requiring case‐by‐case assessments. We present several cases in which variability in results can make clinical decisions complex with limited information and the inability to perform timely confirmatory testing using tests not required by Organ Procurement Transplant Network regulations.

Organ Procurement Transplant Network Policy 2.9.2 requires repeat testing of deceased donors if the donation has not proceeded within 96‐h after the infectious disease screening was initially performed. This study reviews result discrepancies (1.69% discrepancy rate), explores investigations behind peculiar results, and offers several recommendations for managing discrepant results.

Abbreviations

anti‐HBc

Hepatitis B core total antibody

anti‐HCV

Hepatitis C antibody

anti‐HIV

HIV antibody

CDC

Centers for Disease Control and Prevention

CMIA

chemiluminescent microparticle immunoassay

EIA

enzyme‐linked immunoassay

FDA

Food & Drug Administration

HBsAg

Hepatitis B surface antigen

HBV

Hepatitis B

HCV

Hepatitis C

HIV

human immunodeficiency virus

LIMS

Laboratory Information Management System

NAT

nucleic acid test

OPOs

Organ Procurement Organizations

OPTN

Organ Procurement Transplant Network

PHS

U.S. Public Health Service

RR, Non‐D

repeatedly reactive, non‐discriminated

1. Introduction

In March 2021, a new policy was implemented by the Organ Procurement Transplant Network (OPTN) Board of Directors to align with a U.S. Public Health Service (PHS) Guideline for assessing solid organ donors for acute human immunodeficiency virus (HIV), Hepatitis B (HBV), and Hepatitis C (HCV) infection [1, 2]. This policy required Organ Procurement Organizations (OPOs) to draw a repeat blood sample for deceased organ donors if donation had not proceeded within 96‐h after the initial blood sample for screening was obtained [3]. The intent of this policy was to capture potential window period infections that may have gone undetected prior to transplantation causing unintended harm to organ recipients and to maintain safe transplantations.

OPTN Policy 2.9.2 requires donor samples for all required HIV, HBV, and HCV testing must be obtained within 96‐ h prior to organ procurement [3]. Infectious disease testing for all potential deceased organ donors must be executed using FDA‐licensed, approved, or cleared tests for donor screening or diagnostic tests, if no screening test exists for both serology and nucleic acid testing (NAT).

The window period for a test designed to detect a specific pathogen is the time between infection and when the test can reliably detect infection. In antibody‐based testing, the window period is dependent on the time for seroconversion. The concern for transmission of viruses during the window period was a factor in the OPTN policy change. The window period is important because during this time, infection cannot be detected but donors may be able to infect recipients. The advent of NAT decreased the window period for HIV, HBV, and HCV. HIV NAT decreased the window period to 6–11 days compared to 17–22 days with serology testing [4]. The window period for HCV NAT is 3–5 days compared to 70 days for serological tests. HBV NAT window period is 20–22 days versus 35–44 days for antibody tests. The risk of window‐period infection remains exceptionally low, and implementation of NAT has resulted in decreasing the risk of window‐period infection by an order of magnitude or more [5].

During the public comment period for this policy change, the OPO community expressed concerns about the impact of organ allocation [6]. Issues included: repeated testing with new reactive/positive results would result in the need to re‐run matches which could potentially lead to a decrease in the number of organs transplanted and costs associated with additional testing; repeating testing could lead to interpretation/allocation issues if the results do not agree; and lack of clarity on how to manage discordant results. Furthermore, it was recommended that the Disease and Transmission Advisory Committee should consider policy language/guidance regarding repeat testing and guidance when encountering conflicting results.

This retrospective study was performed to determine the frequency in which discrepant results occur with repeated testing recognizing that published guidance lacks for how to interpret and next steps for organ allocation. With over 3 years of experience with this process change, there is an opportunity to develop further guidance for the community to aid in the allocation of donors when discrepant infectious disease results are received.

2. Methods

The infectious disease results from 2653 deceased organ donors were analyzed retrospectively for this study. Donor specimens were collected and tested throughout the United States and tested within 11 different laboratories. Result data was collected from laboratories within a network owned by a single organization that supports multiple OPOs across the country. Donor specimens were submitted to the laboratories from 25 OPOs spanning the Nation. Food & Drug Administration (FDA) approved donor screening tests were used to determine donor eligibility. Hepatitis B core Total and HCV antibodies were detected using either Ortho manual assays or Abbott Alinity s platform and assays. HIV‐1/2 Plus O antibodies and Hepatitis B surface antigen (HBsAg) were detected using either BioRad manual assays or Abbott Alinity s platform and assay. NAT was performed using the Procleix Ultrio Elite assay (Grifols).

Additional testing was performed on some donor cases, post‐organ allocation. Additional testing for HIV utilized the GS HIV‐1 Western Blot (Bio‐Rad) or the Geenius HIV 1/2 Supplemental Assay (BioRad) assays. The GS HIV‐1 Western Blot Kit is an in vitro qualitative assay for the detection and identification of antibodies to HIV‐1. The Geenius Assay is an immunochromatographic assay for the confirmation and differentiation of individual antibodies to HIV‐1 and HIV‐2. The GS HBsAg Confirmatory Assay 3.0 (BioRad) is for the confirmatory neutralization of Hepatitis B surface antigen Reactive specimens. It relies on a human anti‐HBsAg for specificity.

All 11 laboratories were certified by CLIA and held FDA registrations, and each laboratory was audited on a routine basis by these agencies. The laboratories shared SOPs that were housed in an electronic document control system. The SOPs were written verbatim from manufacturer package inserts for all tests. Per the FDA, test protocols cannot be modified, changed, or altered. Quality control was built into each test run and results are documented in the Laboratory Information Management System (LIMS). In addition, the LIMS was programmed to review the QC criteria and result values to indicate a passing or failing test run. Assay sensitivity and specificity were not compared between methods as they are not comparable; NATs detect viral load and serology testing identifies circulating antibodies by enzyme‐linked immunoassays (EIAs, BioRad and Ortho assays) or chemiluminescent microparticle immunoassays (CMIA, Abbott Alinity s platform assays). Comparisons were performed on instrumentation (automated testing platforms, plate washers, plate readers) in instances in which laboratories have platform redundancies. These comparisons took place on a semi‐annual basis to ensure that results were identical despite which instrument was being used. All manufacturer preventative maintenance for instrumentation was performed as prescribed and documented. All laboratories participated in Proficiency Testing surveys from external agencies to demonstrate competency in all testing areas.

Automated CMIA technology was introduced into four of eleven laboratories at different time points during this retrospective study. Donor results were evaluated for increases in discrepancies due to the induction of a new kit vendor and platform and no impact was identified.

Results were queried using data from the laboratories’ Laboratory Information Management System between March 2021 and October 2024.

3. Results

3.1. Discrepancy Rate of Results Obtained From 2653 Deceased Organ Donor Cases

Overall, there was a 1.69% result discrepancy rate (n = 45) comparing results from repeat blood draw specimens to original specimen results. The types of result discrepancies are listed in Table 1.

TABLE 1.

Discrepancy rates by result and by test.

	Discrepancy by result †		% Discrepancy
Result	Reactive to Non‐reactive		0.68
	Non‐reactive to reactive		0.75
	NAT Non‐discriminated to Reactive/Non‐reactive		0.26
	Discrepancy by target
Screening Test	EIA testing	HepB core Total Ab (anti‐HBc)	0.68
		HCV Ab (anti‐HCV)	0.18
		HepB surface Ag	0.07
		HIV1/2 Plus O Ab (anti‐HIV)	0.04
	Nucleic acid testing ‡	HIVd	0.04
		HBVd	0.38
		HCVd	0.04
		Repeatedly Reactive, Non‐discriminated §	0.26

^† Percentage includes both serology and NAT results.

^‡ HIV^d, HBV^d, and HCV^d represent discriminatory testing after a Reactive Ultrio Screen result was obtained.

^§ Reactive NAT Screen followed by Non‐Reactive Discriminatory testing for HIV, HBV, and a second Reactive NAT Screen testing the same draw date/time.

3.2. Discrepancies by Test

Table 1 lists the discrepancy rates for the required HIV, HCV, and HBV donor screening tests. The tests with the highest percentage of result discrepancies were the HBV NAT (0.38%) and Hepatitis B core total (IgM + IgG) antibody (0.68%).

3.2.1. Antibody (Serology) Discrepancies

The Hepatitis B virus core total antibody (anti‐HBc) assay had the highest discrepancy rate for serology testing (0.68%; n = 18. Refer to Table 1). None of these donors were Reactive for HBV NAT and it is probable that false Positive antibody results were obtained (Figure 1). Per the CDC, these results should be interpreted as either (i) acute resolving infection; “window period” if anti‐HBV surface antigen result is negative, (ii) false‐positive total anti‐HBc result, or (iii) recovered from past infection and immune [6].

FIGURE 1.

Discrepant HBc cases. ^†NR = Non‐reactive ^‡R = Reactive.

HBsAg testing had a 0.07% discrepancy rate (n = 2). In one case the result changed from Non‐Reactive to Reactive . The NAT and anti‐HBc results were Non‐Reactive. This result may be explained by recent HBV vaccination in which the surface antigen is transiently positive or a false positive result (refer to the Confirmatory Testing section for further discussion on this donor case) [6]. In another case, both the HBsAg and the anti‐HBc results changed from Non‐Reactive (original result) to Reactive (second blood draw result) ; the donor was NAT Non‐Reactive. The interpretation of the HBV tests on the original specimen would be that the donor is Negative for HBV however, the interpretation for the second specimen test results, per the CDC, the donor recovered from past infection and was immune [6]. This case highlights how there can be confusion with interpreting multiple sets of results in a STAT situation.

HIV‐1/2 Plus O antibody (anti‐HIV) testing had a 0.04% discrepancy rate (n = 1). A Reactive original result was obtained along with a Non‐Reactive result for the second specimen . The HIV NAT results were Non‐Reactive.

HCV antibody (anti‐HCV) testing had a 0.18% discrepancy rate (n = 5). In two cases, Non‐Reactive results were issued for the original blood draw and Reactive for the second blood draw and only one case was Reactive for HCV NAT, indicating that the original anti‐HCV antibody result was a False Negative. For the second case, it cannot be determined if the anti‐HCV result was a False Negative/Positive because people who previously cleared an HCV infection can be NAT Negative and anti‐HCV Positive [7]. Three donors had discrepant anti‐HCV results in which the original results were Reactive, and Non‐Reactive results were obtained when the repeat specimen draw was tested . The HCV NAT results were Non‐Reactive for 2/3 of these donors. People previously infected and have cleared the viral infection (evidenced by NAT Negative results), can have detectable anti‐HCV. For the one donor in which NAT was Reactive, the original anti‐HCV result was Reactive and the second blood draw was Non‐Reactive , it is likely that the latter results were False Negative.

3.2.2. NAT Discrepancies

Ultrio NAT Screen and Discriminatory result discrepancies account for 0.72% of the repeat sample discrepancies. Discrepancies are presented in Table 1. The Ultrio HBV NAT Discriminatory discrepancy rate was 0.38% (n = 10) and 0.04% for both HIV and HCV NAT (n = 1, for each virus).

On occasion (0.26% of all donors tested), an Ultrio NAT Screening result cannot be obtained, and the testing algorithm yields a result of repeatedly reactive, non‐discriminated (RR, Non‐D); refer to Figure 2 for an outline of the complex testing algorithm [9]. Non‐reactive HIV, HBV, and HCV results from second blood draw specimens were obtained for 5/7 donors that originally had Repeatedly Reactive, Non‐Discriminated results. Serology results were varied for these donors (refer to Table 2) however, likely False Positive antibody results were obtained and organs were able to be placed.

FIGURE 2.

Nucleic acid test algorithm and result discrepancies encountered during testing. The number of discrepancies is marked by circled numbers. Results that changed from Non‐Reactive to Reactive for either human immunodeficiency virus (HIV), Hepatitis B (HBV), or Hepatitis C (HCV) are noted by Results that changed from Reactive to Non‐reactive are noted by. Results that changed from Repeatedly Reactive and Non‐discriminated to Non‐reactive are noted by.

TABLE 2.

Occasionally, nucleic acid test (NAT) results do not yield informative clinical information and upon testing of the second blood draw, produced useful results for allocation.

Non‐discriminated NAT (Original result)/Non‐reactive (Second draw result)
Examples	Anti‐HIV‐1/2 Plus O	Anti‐HCV	Anti‐HBc	HBsAg	Comments	Allocation status
1	NR	NR	R	NR	Potential False Pos anti‐HBc †	HBc reactive/NAT NR Kidneys and liver placed
2	NR	NR	R	NR	Potential False Pos anti‐HBc ‡	HBc reactive/NAT NR Heart, liver, and lungs placed
3	NR	R	NR	NR	HCV NAT Non‐reactive ‡	HCV reactive NAT NR Kidneys placed
4	NR	NR	R	NR	Potential False Pos anti‐HBc †	HBc reactive/NAT NR Liver and heart placed
5	R	NR	NR	NR	Potential False Negative NAT §	Non‐donor due to other factors.

^† refer to CDC Table 3‐1. Interpretation of hepatitis B laboratory results.

^‡ anti‐HCV testing was not performed on the 96‐h specimen. Cannot determine if the original result was a False Positive.

^§ Western Blot or confirmatory antibody testing was performed.

3.3. Additional Testing

Additional testing, or testing performed outside of the OPTN requirements, can be useful to gain an understanding of conflicting test results or can be used to counsel donor families when unexpected screening results are obtained. In one case, the donor was Reactive for HIV antibodies upon initial testing and Non‐Reactive when the second blood draw specimen was tested. NAT results were non‐reactive and confirmatory HIV 1/2 antibody testing yielded Non‐Reactive results indicating that the original serology result was a False Positive. This donor was allocated as HIV Negative and all results were shared with the transplant centers during placement of organs.

We noted another case in which a false positive result was obtained as confirmed with a confirmatory test. The second draw specimen was Reactive for HBsAg. The surface antigen neutralization assay was performed, and the result was Not Detected. Per the CDC, for HBV tests, to resolve the ambiguity of a false‐positive total anti‐HBc result, a follow‐up sample drawn 4–8 weeks later should be tested and if the result is Negative, interpreted as false‐positive [7]. In the setting of deceased organ donation recovery, this type of re‐test is not a possibility.

A False Negative HIV NAT result was obtained for one donor. The second blood draw specimen was HIV NAT Reactive while the original specimen was NAT Non‐Reactive. The HIV antibody results were Reactive for both specimens. Furthermore, post‐allocation, HIV Western Blot testing was performed using both specimens and the results were positive. The initial Ultrio Screen result for this donor can be considered a False Negative. It has been previously reported that 18% of donors that were serologically Reactive for anti‐HIV and HIV NAT non‐reactive had positive HIV western blot results [10]. Western Blot testing was performed after allocation to help make sense of the HIV serology results and hence the determination of false negative NAT results.

3.4. Transfusion Status

We investigated whether transfusion status may play a role in explaining some of the result discrepancies. For example, three [3] donor cases had discrepant HBV NAT results. Reactive HBV NAT results were obtained when the second blood draw specimens were tested. The original specimens were drawn pre‐transfusion and the second blood draw specimens were drawn post‐transfusion. Adding in a layer of uncertainty, the OPOs ordered testing for a third time for these donors, post‐transfusion, and Non‐Reactive HBV NAT results were obtained, bringing into question the true infectious status of the donors. The NAT and serology results were varied which can cause confusion at the time of allocation.

We categorized cases with discrepant results in which donors had been transfused. On occasion, the original specimens tested were pre‐transfusion draws and the second blood draws were post‐transfusion (0.43% of all donors tested), and in even rarer instances both blood draws were post‐transfusion specimens (0.20%). There was no apparent link between discrepant results and donor transfusion status.

3.5. Laboratory Factors

There was no evidence of clustered discrepant results within a close timeframe which could indicate errors in testing (personnel or equipment). There were no indications of issues with assay manufacturer lots. During the course of this study, four [4] laboratories implemented an automated serology platform, Alinity s (Abbott) System. We evaluated whether the introduction of the Alinity s increased or decreased the amount of discrepant serology results. There is no correlation between manual versus automated serology testing that contributed to discrepant results.

Discrepant results by laboratory location were reviewed. The laboratories were categorized for High, Medium, or Low STAT volumes during the timeframe of this study (all donors tested at each lab factored into the designated volume of each laboratory, not just the donors that were tested on two occasions). There was a moderate correlation between the donor volume of the second blood draw specimens tested and the discrepancy rate (r = 0.55). There is no variation between these laboratories due to the consistent quality control measures implemented at each site (refer to the Methods section for further details). Additionally, all technologists followed the same training protocol and performed testing using identical standard operating procedures. As a result, no variability has been introduced in this study due to geographic differences.

4. Discussion

The intent of revising the OPTN policy was to maintain transplant recipient safety while more accurately identifying organ donors that have certain risk factors for acute HIV, HBV, and HCV infection. This data represents that there is more than a 1% chance that discrepant results will be obtained. The application of this new policy has introduced complexity, confusion, and delays in the allocation process when discrepant results are obtained, especially given the lack of any formal community guidance in such instances. We have presented several cases in which variability in results can make clinical decisions complex with limited information and the inability to perform timely confirmatory testing using tests not required by the OPTN regulations. Considerations should be made to revise the requirement to only apply to cases where the potential donor has a known history of intravenous drug abuse or other high‐risk behavioral risk factors. Considerations should be made to provide clarity to the community on how to document and declare organ donor results as well as consistency in how to determine those results.

We estimate from the review of thousands of donor screening cases, that 0.70% of discrepant results can be explained by False Positive results, either initially Reactive results and Non‐Reactive 96‐h specimen results or vice versa. The False Negative rate was estimated at 0.1%. A limitation of this retrospective review is that confirmatory testing was not conducted on discrepant donor results to determine true false positivity rates. Excess specimens are preserved and entered into long‐term storage should the need arise for additional testing in the event of suspected disease transmission. As such, specimens are not always available for testing for research purposes. As the occurrence rate for discrepancies is low, it is in these rare circumstances that confusion can occur, thereby delaying decisions while the OPO teams determine the path ahead. Of importance to note, if for example, an HBV NAT Reactive result is obtained on the original blood specimen, the 96‐h rule would indicate that HIV and HCV NAT testing is required. To test for these two viruses, per manufacturer package insert, the Ultrio screening assay (screens for all three viruses at one time) must be performed prior to discriminatory NAT testing, and only if the Ultrio screen is Reactive can the discrimination NAT occur. The discrimination testing that must be performed is for all three viruses. Herein lies a small chance of receiving discrepant results. Thirdly, attempts at gaining a NAT result in instances in which repeatedly reactive, non‐discriminated results are obtained should be considered. Despite the low number of instances observed in the data cohort, one hundred percent of these donors ultimately had Non‐Reactive NAT results with further testing.

It is not clear what causes discrepant results. These assays were initially intended for screening donors for blood donation and have not been widely studied with blood from deceased organ donors. Some manufacture package inserts state that reactivity at or slightly above the cutoff value is more frequently nonspecific, especially in specimens obtained from persons at low risk for infection [11, 12]. Another vendor published that false reactive results can be expected with any test kit and may be observed due to non‐specific interactions [13]. This may explain why some Reactive antibody results do not correlate with non‐reactive NAT results. Abbott performed a study looking at interference in the anti‐Hepatitis B core total assay with total Bilirubin (≤ 20 mg/dL), unconjugated Bilirubin (≤ 20 mg/dL), hemoglobin (≤ 500 mg/dL), triglycerides (≤ 3000 mg/dL), and total Protein (≤ 12 g/dL) and found no interference [13]. No other substances were investigated. General chemistry results for deceased donors do not always fall within the range of Normal and it is difficult to predict how these results may interfere with donor screening assays for infectious diseases.

Discrepant results trigger case‐by‐case assessments that should include consultations with ID specialists and/or Chief Medical Officers. OPOs need to evaluate risk as well as all available results. OPOs are required to disclose all results and should ensure clear documentation clarifying testing discrepancies so transplant centers can make their own informed assessments. While it is not possible to correlate discrepant result instances with organ discard rates, there are clinical consequences of result discrepancies that impact organ allocation. There is an impact on the organ allocation process by introducing discordant results after primary allocation and therefore creating inefficiencies in the system. Additionally, transplant centers need to interpret the discordant results in the absence of formal guidance. The downstream effect of this is an adverse impact on patient selection given that the original Negative test result is now Positive; donors may be headed to surgery or already in the operating room resulting in less time for transplant programs to identify appropriate patients. There may also be further deleterious effects ultimately causing organ non‐use, due to these confounding factors.

There is an opportunity to consider the rationale for the requirement for repeat serology (antibody) testing, post‐96‐h for HBV, HCV, and HIV given the unlikelihood that a patient would seroconvert antibody with negative NAT. The intent of a second blood draw resting is to reduce the likelihood of overlooking emerging risks. Employing NAT addresses this concern. NAT methodologies are more sensitive than antibody testing and will detect infection sooner.

Policy changes should be routinely evaluated for their effectiveness, with a particular focus on establishing community guidelines for the disclosure and interpretation of conflicting results. Additionally, careful consideration of cost and time implications is essential to ensure effective stewardship in the deceased organ donor evaluation process. Revisions to current requirements—such as restricting certain protocols to cases where the potential donor has a known history of high‐risk behaviors and has not already tested positive for NAT or antibodies during donor screening—merit further consideration. Despite these complexities, the risk of disease transmission through organ donation remains low, due to the use of sensitive molecular testing [14]. Testing a second sample by NAT minimizes the risk of transmitting undetected infections to recipients.

Based on this data, we offer several recommendations for managing discrepant results. First, OPO staff should be educated on the potential for discrepancies, emphasizing that while rare, they can occur and are not always attributable to laboratory error. Second, OPOs should establish a defined protocol for addressing discrepant results to ensure efficiency when managing organ allocation, while transplant centers must have a plan for recipient management under such circumstances. For instance, if a donor is anti‐HBc Positive and HBV NAT Neg, per the CDC, this may represent a false‐positive total anti‐HBc result. To resolve this ambiguity, a follow‐up sample should ideally be tested 4–8 weeks later. If the follow‐up test remains positive, it should be interpreted as evidence of a resolved infection; if negative, the initial result can be considered a false‐positive anti‐HBc result. However, due to the time‐sensitive nature of the deceased donor process, repeating tests weeks later is not feasible. This underscores the critical importance of understanding test results—particularly in the context of discrepant findings—so that organ procurement and transplant teams can make informed decisions and maximize the number of organs available for transplant.

Conflicts of Interest

Sean Van Slyck serves as the Executive Director for Sierra Donor Services and is the co‐Chair of the Scientific Registry of Transplant Recipients Review Committee. Chris Curran serves as the Senior Vice President of Organ Utilization for New England Donor Services and serves on the OPTN Membership Professional Standards Committee. There are no conflicts of interest for all the authors and no financial support was received for this study.

Ethics Statement

Ethical approval was not sought for the present study because the data presented here was collected retrospectively; consent of donors occurred prior to donation for screening of infectious diseases as part of the routine donation process. This data is not sensitive and has no risk of compromising the identity/safety of patients.

Dionne S., Akana H., Curran C., and Van Slyck S., “Evaluation of Discrepant Infectious Disease Results in Deceased Organ Donors: Insights From a Retrospective Analysis of Post‐Policy Testing.” Transplant Infectious Disease 27, no. 4 (2025): 27, e70055. 10.1111/tid.70055