Human CLDN1/Claudin-1 ELISA Kit

A sandwich ELISA using a double antibody configuration, the Human CLDN1/Claudin-1 ELISA Kit from Assay Genie is designed for the quantitative measurement of CLDN1 levels in human samples. Detection is colorimetric with measurement at 450 nm following the addition of stop solution, and the kit is stored at 2-8°C for 12 months. CLDN1, also known as Claudin-1, is a protein involved in the regulation of tight junctions between cells, contributing to epithelial barrier function and cell polarity, with dysregulation implicated in cancer, inflammatory bowel disease, and hepatitis.

The assay begins with equilibration of reagents and TMB substrate to room temperature, followed by loading of standards, samples and control (zero) wells onto the pre-coated plate and incubation at 37°C for 90 minutes. A biotin-labeled detection antibody is added and incubated for 60 minutes at 37°C, then HRP-Streptavidin (SABC) is added for 30 minutes at 37°C. After TMB substrate is added and incubated in the dark for 10–20 minutes, stop solution is added and absorbance is read immediately at 450 nm.

Sample preparation protocols include allowing blood to clot and centrifuging at 1000 × g for 20 minutes for serum; collecting plasma using anticoagulant tubes and centrifuging at 1000 × g for 15 minutes at 2–8°C; homogenising tissue in PBS with protease inhibitors and collecting the supernatant; centrifuging cell culture supernatant at 2500 rpm for 5 minutes; and lysing cells with lysis buffer containing protease inhibitors. Linearity in serum (n=5) was 86-95% at 1:2, 80-94% at 1:4 and 90-103% at 1:8; EDTA plasma (n=5) showed 83-98%, 81-100% and 87-105%; and heparin plasma (n=5) showed 86-101%, 84-100% and 85-94%. Recovery ranges were 93-103% with an average of 98% for serum, 89-103% with an average of 92% for EDTA plasma, and 92-105% with an average of 98% for heparin plasma.