Sample‐to‐cutoff ratios using Architect HIV Ag/Ab Combo: The influence with the results of supplemental tests and optimal cutoff value to predict HIV infection

Background The Architect HIV Ag/Ab Combo has excellent performance for HIV screening; however, the false‐positive rate (FPR) was high in low HIV prevalence setting. Objectives The purpose of this study was to analyze the influence of sample‐to‐cutoff (s/co) ratios by Architect HIV Ag/Ab Combo with the results of confirmatory test and explore the potential utility of s/co to predict HIV infection. Methods A retrospective review on Architect HIV Ag/Ab Combo reactive results was performed at a teaching hospital in Xi'an. The s/co values in different groups, that is, true positives (TP) and false positives (FP), different Western blotting (WB) bands among WB‐positive cases, were compared. The receiver operating characteristic curve (ROC) analysis was used to determine the optimal cutoff value for predicting HIV infection. Results During the study period, 219 out of 84 702 patients were reactive by ARCHITECT with a 0.0992% of HIV prevalence and a 56.25% of FPR. The mean s/co ratios in TP were significantly higher than that in FP (458.15 vs 3.11, P < 0.0001). Among the WB‐positive cases, the s/co ratios increased significantly with the increase in the number of bands, P = 0.0065. The optimal cutoff (24.44) by ROC analysis can provide the highest sum of sensitivity (100%) and specificity (100%) with no FP results. Conclusions For Architect HIV Ag/Ab Combo, the FPR is reduced when s/co ratios increase, and the s/co ≥24.44 may be reliable to predict HIV infection.


| BACKG ROU N D
HIV/AIDS is a serious public health issue. Data from World Health Organization (WHO) have shown that people living with HIV (PLWH) were more than 35 million and 940 000 people died from HIV-related causes globally in 2017. 1 Since the HIV/AIDS was first reported in Beijing and Zhejiang Province in 1985, [2][3][4] the spread rate of HIV in China is alarming. Quite a few prevention policies and tremendous efforts, such as ban on imported blood products since 1985, 3 blood donors must be tested for HIV since 1995, methadone maintenance treatment program and four free one care, 2 The ARCHITECT HIV Ag/Ab Combo, which based on a chemiluminescent immunoassay with the ability to simultaneously detect p24 antigen and HIV-1/2 antibodies, has shortened the window period by around 1 week compared to the third-generation assay [9][10][11] and has a broader dynamic readout range. However, the rate of false positives (FP) by the fourth-generation assay was high in situations of low HIV prevalence. [12][13][14] The present study was carried out to analyze the influence of s/co ratios by Architect HIV Ag/Ab Combo with the results of supplemental tests and explore the optimal cutoff value to predict HIV infection.

| Two-step HIV diagnostic algorithm
The nucleic acid and p24 antigen tests were not applied at Xi'an CDC; meanwhile, low HIV prevalence remained in China and positive predictive value of the fourth-generation assay was poor. [15][16][17][18] Considering that Western blotting (WB) is a confirmatory test, a WB-negative result is often regarded as exclusion of HIV infection by the patient and clinician in China. The incorrect understand and treatment may lead to serious consequences for the early infections.
Since January 16, 2017, a new delivery protocol of blood samples that only reactive to HIV-1/2 Ab subjects should be conducted and WB has been developed by Xi'an CDC.
Based on this, a two-step HIV diagnostic algorithm is currently used at our hospital. The first step, a fourth-generation kit, Architect HIV Ag/Ab Combo (Abbott Diagnostics, Abbott Park, IL), which allows for identification of acute infection and importantly reduces the antibodyfree "window" period, is used as the HIV-1/2 initially screening test.
The second step, a third-generation EIA kit, XinChuang HIV-1/2 Ab (InTec, INC, XiaMen, FuJian, China), and a confirmatory test (WB) are used as the supplemental tests for the initially screening reactive cases.
For the third-and fourth-generation kits, the s/co ≥1 and s/co <1 were defined as reactive and non-reactive (negative), respectively.
HIV-1/2 antibodies and p24 antigen negative should be reported if the initially screening test is non-reactive. WB should be conducted if it was reactive to HIV-1/2 Ab. For the only HIV Ag/Ab Combo reactive subjects, the HIV-1/2 Ab test should be carried out at the 2nd, 4th week, 3rd and 6th month, and the WB should be performed once HIV-1/2 Ab reactive occurs. For WB-negative or indeterminate cases, WB should be conducted at the 2nd, 4th week, 3rd and 6th month.

| Western blotting
Western blotting (HIV1/2 BLOT 2.2; MP Biomedicals, Singapore) is conducted at the Xi'an CDC, with the separated HIV-1 gene product

| Statistical analysis
Statistical analyses were conducted by SPSS13.0 (serial number 5026743; SPSS Inc, Chicago, IL), and WB positive was the standard for HIV infection diagnosis in the study. The continuous variables were expressed by mean, the Mann-Whitney U test was used between two groups, and multiple groups were performed using the one-way ANOVA test and Kruskal-Wallis test. The receiver operating characteristic curve (ROC) analysis was used to determine the optimal cutoff value for predicting true positives (TP). A P-value <0.05 (two-tailed) was considered as statistically significant.

| Analysis of Western blotting results
Among the 69 WB-positive cases, there were 15 kinds of WB patterns, 84.06% (58/69) cases presented with reactivity to more than or equal to seven antigens patterns, and the s/co ratios had a statistically significant association with the band numbers, P = 0.0065.
According to the follow-up results of the WB-negative and indeter-

ACK N OWLED G M ENTS
The authors would like to thank Hai-Chao Zheng from Xi'an CDC, Shaanxi, China, for his contribution of the data collection in the study.

AUTH O R S' CO NTR I B UTI O N S
LW and YY were major contributors in the writing of the article. LW, JYW, and FF were responsible for the study design. The statistical analysis and figure of the study were performed by LW, JYW, JXR, XDT, and YY.

E TH I C S S TATEM ENT
The study was deemed exempt from review by the Ethics Committee of the First Affiliated Hospital of Xi'an Jiaotong University as routine data for clinical purpose were used and all the information of patients was kept confidential in the study.

CO N S E NT TO PA RTI CI PATE
Not applicable.

CO N S E NT FO R PU B LI C ATI O N
LW, JYW, JXR, XDT, YY, and FF have read and approved the final work for publication in Clinica Chimica Acta.

AVA I L A B I LIT Y O F DATA A N D M ATE R I A L
The data used in the study were available from Xi'an CDC and LIS of the First Affiliated Hospital of Xi'an Jiaotong University.