This protocol was already used successfully to detect anti-HIV antibody in the serum of women with cervical dysplasia or cervical cancer in Jamaica, West Indies [1]. This is an open access protocol distributed under the terms of the C rea tiv e C o m m o n s A ttrib u tio n Licen se C rea tiv e C o m m o n s A ttrib u tio n Licen se (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and

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... are credited 1 The 96 well polystyrene microplate (U-shaped bottom; Sigma-Aldrich) is coated with 50 ng of a mixture of synthetic peptides (including the fragment 579-601 of the HIV gp41 and fragments 254-274, 308-331 and 421-438 of the HIV gp120) for 4 h at 37ºC. 2 The microplate is blocked with 3% non-fat milk in PBS, 25 µl/well, 1h at room temperature (RT). 3 The microplate is washed 4X with PBS-Tween-20. 4 Duplicates of 25 µl of 1:16 diluted human sera are added. 5 After incubation for 90 min at RT the microplate is washed 4X with PBS-Tween 20. 6 Then, 25 µl of a chimeric commercially-prepared recombinant protein LA-HRP conjugate (Sigma-Aldrich) diluted 1:5000 is added. 7 After incubation for 90 min at RT and rewashing steps 25 µl TMB is added to each well for 15 min in the dark. 8 The reaction is stopped with 3M H2SO4. 9 The microplate is read in a microplate reader at 450 nm. 10 In the ELISA is included a pooled human sera with high titre of anti-HIV antibodies as positive control, a pooled sera from healthy individuals as negative control and 0.9% normal saline solution was used as the blank. 11 The cut-off point is calculated as mean optical density (XOD) of negative control plus two standard deviation (SD). This is an open access protocol distributed under the terms of the C rea tiv e C o m m o n s A ttrib u tio n Licen se C rea tiv e C o m m o n s A ttrib u tio n Licen se (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited