Monoclonal rabbit anti phospho p44 42 mapk erk1 2 thr202 tyr204 antibody

Levels of the biomarker p-ERK in the excised cNFs were defined as the p-ERK proportion of total ERK in the tumor protein lysate using a size-based nano-immunoassay (the Peggy Sue platform, Protein Simple). Antibodies used included monoclonal rabbit anti–phospho-p44/42 MAPK (Erk1/2) (Thr²⁰²/Tyr²⁰⁴) antibody (Cell Signaling Technology, catalog no. 4370L) and monoclonal rabbit anti-p44/42 MAPK (Erk1/2) antibody (Cell Signaling Technology, catalog no. 4695S). Target tumor diameter and height measured by ruler were the basis of assessments of tumor volume. Volume was calculated on the basis of ruler measurements of longest diameter and height using a calculation of volume as a cylinder with known diameter and height. For systemic pharmacokinetic analysis, samples were collected at one site for evaluation of plasma levels of NFX-179.

Ex vivo assays were used to determine the activity of NFX-179 Topical Gel in penetrating skin and suppressing the MAPK pathway and to guide dosage choices for the clinical study. NFX-179 Topical Gel 0.1%, 0.5%, or vehicle was applied to the epidermal surface of surgically excised cNF tumors semi-immersed in media with the dermis in media and the surface of the skin exposed for 4 hours (n = 3 for each dose). After drug treatment, the tissue was processed for Western blot and immunohistochemistry analysis of p-ERK, a downstream target of the MAPK pathway. Antibodies used included monoclonal rabbit anti–phospho-p44/42 MAPK (Erk1/2) (Thr²⁰²/Tyr²⁰⁴) antibody (Cell Signaling Technology, catalog no. 4370 L), mouse anti–β actin (Sigma-Aldrich, catalog no. A1978), goat anti-mouse immunoglobulin G (IgG) (Jackson ImmunoResearch), and goat anti-rabbit IgG (Thermo Fisher Scientific).