Summary/Abstract High throughput sequencing of the rearranged T cell receptor genes (HTS) has transformed the diagnosis, care and assessment of therapeutic responses in patients with cutaneous T cell lymphoma (CTCL) and this assay is becoming the gold standard in CTCL clinical trials. HTS results are highly reproducible in frozen CTCL skin biopsies but the formalin used to preserve skin biopsies in many clinical trials degrades DNA and affects HTS measurements, potentially causing errors in patient diagnosis, assessment of responses and choices of therapy. We seek to identify a single uniform tissue processing approach for small CTCL skin biopsies that will give accurate and reproducible HTS results, support DNA, RNA and protein measurements, maintain excellent histology, and preserve remaining tissue for future measurement of emerging biomarkers. In Aim 1, we identify optimal tissue transport conditions and test non-cross-linking fixatives for their ability to support nucleic acid, protein and histologic studies on small skin biopsies. Aim 2 studies the effects of storage time and temperature on nucleic acid integrity, histologic performance and HTS readings, and tests two approaches to mitigate the effects of DNA degradation on HTS measurements. Aim 3 provides real world testing of our optimized sample handling procedures, using them to study skin biopsies obtained in the industry sponsored, randomized, placebo- controlled phase III trial of topical resiquimod gel in CTCL. Our overall goals are to establish new best tissue handling practices for future clinical trials and to establish corrections that allow accurate analyses of existing specimens. HTS is now frequently used in many cancer types to measure tumor T cell numbers, diversity and responses to immune therapies. The optimized tissue handling procedures we identify therefore have the potential to be useful in many cancer types.