Protocol detail

Assessing HPV16-specific T-cell Responses

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Eight peptide pools of HPV16 peptides were used to determine the proliferative capacity of HPV16-specific T-cells, for cytokine polarization analysis and detection of the HPV16-specific Foxp3+ regulatory T-cells, as described previously [10 (link), 21 (link), 22 (link)]. Panels of overlapping 30-35 mer peptides with HPV16 E2, E6, and E7 protein sequences were synthesized by solid phase peptide synthesis (SPPS) method with >95 % purity (ChinaPeptides Co. Shanghai, China), with a 14 (for 30-mer) or 15 (for 35-mer) amino acid (aa) overlap. Two pools of E2 peptides (E2.1 and E2.2) consisted of 12 or 11 (30-mer) peptides, respectively. Four pools of E6 and two pools of E7 peptides (E6.1-E6.4 and E7.1 and E7.2) consisted of two 32-mer or 35-mer peptides, respectively [10 (link)]. The numbers of covered amino acids of each protein are shown in Additional file 1. Memory response mix (MRM) stock solution (50×), consisting of tetanus toxoid, 0.75 fL/mL (Statens Serum Institut, Copenhagen, Denmark), Tuberculin PPD, 5 µg/mL (Statens Serum Institut), and Candida albicans, 0.015 % (Greer Laboratories, Lenoir, USA) was used as a positive control for the proliferation assays and cytokine production capacity of the PBMCs.

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Koskimaa H.M., Paaso A., Welters M.J., Grénman S., Syrjänen K., van der Burg S.H, & Syrjänen S. (2015). Human papillomavirus 16-specific cell-mediated immunity in children born to mothers with incident cervical intraepithelial neoplasia (CIN) and to those constantly HPV negative. Journal of Translational Medicine, 13, 370.

Publication 2015

tetanus toxoid Tuberculin PPD Candida albicans

Amino acids Assays Candida albicans Cytokine Hpv16 Memory Peptides Protein Protein sequences Regulatory t cells Serum Spps T cells Tetanus toxoid Tuberculin ppd

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Variable analysis

independent variables

Eight peptide pools of HPV16 peptides (E2.1, E2.2, E6.1-E6.4, E7.1, E7.2)

dependent variables

Proliferative capacity of HPV16-specific T-cells
Cytokine polarization analysis
Detection of the HPV16-specific Foxp3+ regulatory T-cells

control variables

Memory response mix (MRM) stock solution (50×), consisting of tetanus toxoid, 0.75 fL/mL, Tuberculin PPD, 5 µg/mL, and Candida albicans, 0.015 %

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