Antibody Hybridization Dataset with Cytokeratin Probes and Contact Epithelial Cells

Touch samples were collected from volunteers using the following protocol which was approved by the VCU-IRB (#HM20000454_CR). Contributors either rubbed a sterile polypropylene conical tube (P/N 229421; Celltreat Scientific) for five minutes using their entire hand (i.e., palm and fingers) and held it in their hands (no rubbing) for five minutes. 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).

For antibody hybridization experiments, 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 CK probes. This was allowed to incubate at room temperature for 10 minutes. Cells were then incubated with either cytokeratin probe ‘AE1’ which recognizes CKs 10, 14, 15, 16 and 19 or 'AE3' which recognizes CKs 1, 2, 3, 4, 5, 6, 7, 8), Cat#s 14-9001-80 and 14-900-80, respectively, Affymetrix eBioscience) for 30 minutes followed by reaction with a secondary antibody, anti-mouse IgG1-APC (Cat# 17-4015-80, Affymetrix eBioscience). We used anti-mouse IgG1-APC (Cat#17-4714-42, Affymetrix eBioscience) to create the isotype control for AE1 experiments, incubating for 30 minutes. Cells were washed once and then resuspended in 1xFACS buffer prior to analysis.

Flow cytometry analysis was performed on the BD FACSCanto™ II Analyzer (Becton Dickinson) equipped with 488nm and 633nm lasers. Channel voltages were set as follows: Forward Scatter (FSC, 150V), Side Scatter (SSC, 200V), Alexa Fluor 488 (FITC, 335V), Phycoerythrin (PE, 233V; PE-Cy5, 300V; PE-Cy7, 400V), and Allophycocyanin (APC, 250V).

Flow cytometry source data for all samples are provided in Flow Cytometry Standard (.fcs) format files. Source data files are labeled by date of collection, ID, and the antibody probe used. Isotype and unstained controls are included.