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. 2025 Jan 2;25(1):6.
doi: 10.1186/s12879-024-10404-1.

Modified TPPA combined with western blotting facilitates syphilis diagnosis in isolated reactive treponemal chemiluminescence immunoassay sera: a prospective cohort study

Xia Luo #  1 , Hua Xiao #  2 , Yanfang Lu  3   4   5   6 , Weiming Gu  7 , Ziyong Sun  1 , Jing Peng  8 , Liming Cheng  9
Affiliations

Modified TPPA combined with western blotting facilitates syphilis diagnosis in isolated reactive treponemal chemiluminescence immunoassay sera: a prospective cohort study

Xia Luo et al. BMC Infect Dis. .

Abstract

Background: The challenge of dealing with isolated reactive treponemal chemiluminescence immunoassay (CIA) results in clinical practice has prompted the development of a more efficient algorithm for distinguishing true infection from false reactivity in isolated CIA sera.

Methods: A prospective cohort study was conducted at Wuhan Tongji Hospital, involving 119,002 individuals screened for syphilis using CIA from January 1, 2015, to January 6, 2017. Samples with reactive CIA results underwent simultaneous testing with the T. pallidum passive particle agglutination assay (TPPA) and the rapid plasma reagin test (RPR). Additionally, a subgroup of 189 individuals with differing TPPA statuses was selected for further analysis using Western blotting (WB) and a modified TPPA assay (titer, 1:20). To identify the optimal serological approach for distinguishing true from false reactivity in sera with isolated reactive CIAs (CIA+TPPA-RPR-), two distinct algorithms were developed and evaluated. The first algorithm involved reflexively testing CIA+TPPA-RPR- sera with the modified TPPA, followed by WB if nonreactive. The second algorithm began with WB, followed by the modified TPPA if nonreactive or indeterminate.

Results: WB demonstrated lower sensitivity compared to TPPA, but it identified six syphilis cases among the 89 CIA+TPPA- samples. Both WB and modified TPPA exhibited a specificity of 100%. The two supplementary confirmatory algorithms detected 12 additional syphilis cases, with the first algorithm being more cost-effective and labor-saving.

Conclusion: A combination of a modified TPPA (titer, 1:20) and WB can serve as a reliable algorithm for distinguishing true syphilis infection from false reactive signals in isolated reactive CIA sera.

Clinical trial number: Not applicable.

Keywords: Isolated reactive chemiluminescence immunoassay; Modified TPPA (titer, 1:20); Syphilis serodiagnosis; Western blotting.

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Conflict of interest statement

Declarations. Ethics approval and consent to participate: This study was conducted in accordance with the principles outlined in the Declaration of Helsinki and was approved by the Institutional Review Board of Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology (TJ-IRB20231172). The requirement for consent to participate was waived by the Institutional Review Board of Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, as the study involved the use of de-identified medical records and residual serum for secondary analysis, with no direct participant intervention. Consent for publication: Not Applicable. Competing interests: The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
Distribution of CIA signal/cutoff values in different serum groups. (A) Signal/cutoff value distribution in CIA+TPPA−RPR− (n = 478), CIA+TPPA+RPR− (n = 1034) and CIA+TPPA+RPR+ (n = 791) groups. (B) RPR titers versus signal/cutoff values in CIA+TPPA+RPR+ sera. +, reactive; −, nonreactive; RPR (titer, 1:1), n = 477; RPR (titer, 1:2), n = 128; RPR (titer, 1:4), n = 76; RPR (titer, 1:8), n = 33; RPR (titer, 1:16), n = 34; RPR (titer, 1:32), n = 19; RPR (titer, > 1:32), n = 24. Abbreviations: CIA, chemiluminescence immunoassay; TPPA, Treponema pallidum particle agglutination assay; RPR, rapid plasma reagin test
Fig. 2
Fig. 2
TPPA compared to WB as a confirmatory test for syphilis. (A) WB results of samples with CIA+TPPA+ (n = 61). (B) WB results of samples with CIA+TPPA− (n = 89). (C) WB results of samples with CIA−TPPA− (n = 39). Abbreviations: TPPA, Treponema pallidum particle agglutination assay; WB, Western blotting; +, reactive; −, nonreactive; ±, indeterminate
Fig. 3
Fig. 3
Working algorithms for isolated reactive CIA sera. (A) The algorithm starting with the modified TPPA (titer, 1:20). (B) The algorithm starting with WB. Abbreviations: TPPA, Treponema pallidum particle agglutination assay; RPR, rapid plasma reagin test; WB, Western blotting; +, reactive; −, nonreactive; ±, indeterminate
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