Antibody Hybridization Dataset Using Human Leukocyte Antigen (HLA) Probes and Epithelial Cells (Buccal and Contact Epidermal)

Biological samples were collected from volunteers using the following protocol approved by the VCU-IRB (#HM20000454_CR). For contact epithelial samples contributors rubbed a sterile polypropylene conical tube (P/N 229421; Celltreat Scientific) for five minutes using their entire hand (i.e., palm and fingers). Cells were then collected from the surface with six sterile pre-wetted swabs (P/N 22037924; Fisher Scientific) followed by two dry swabs. To elute the cells into solution, the swabs were manually stirred then vortexed for 15 seconds in 10 mL of ultrapure water (18.2 MΩ∙cm). Buccal cells were collected by swabbing the inner cheek surface with a cotton-tipped swab. As before, swabs were mixed in 10mL of ultrapure water to create a cell solution used for subsequent hybridization experiments. 

For antibody hybridizations, three milliliters of each cell solution were centrifuged at 5,000xg for five minutes. The supernatant was decanted and the pellet was resuspended in 100 µl PBS buffer and 1 µL of Human Fc Receptor block (Cat# 130-059-901, Miltenyi Biotec) to increase the specificity of antibody binding before reaction with HLA probes. This was allowed to incubate at room temperature for 10 minutes. Cells were incubated with mouse anti-human monoclonal antibody (mAb) HLA-ABC-FITC (Cat# 311403, BioLegend) or HLA-A02 (Cat#343303, Biolegend)  for 30 minutes.  Cells incubated with anti-mouse IgG2a-FITC (Cat# 343303, BioLegend) for 30 minutes served as the isotype control for these experiments. Cells were then washed once in 1x FACS buffer [PBS supplemented with 2% Fetal Bovine Serum (FBS, Cat# 100-106, Gemini BioProducts) and 10% Sodium Azide (Cat# S2002, Sigma-Aldrich)] and re-suspended in the same solution until flow cytometry analysis.