Representative Western blot analysis for the selected panel of differentially expressed targets in the various T cell subsets.

Western blot analyses were performed by serially probing sets of 3 membranes per donor. The expression levels of GDIR1, GSTO1, LIMS1 and PROF1 were analyzed blot 1 (upper panel), GDIR2, PRDX2 and THIO on blot 2 (middle panel) and SODM on blot 3 (lower level) of the respective membrane set.

The relative expression levels were defined by densitometric quantification normalized to the corresponding beta-actin signals, co-detected on each membrane.

Aliquots of 15 µg total lysate per sample/lane were loaded onto Tris-Tricine gels (16%T/2.5%C) and subsequently separated. Lysates generated form untreated CD45RA⁺ and CD45RO⁺ T cells were throughout the set of membranes loaded onto the lanes 1 and 3 respectively, whereas lysates generated from the corresponding T cell subsets exposure for 3 h to 5 µM H₂O₂ were loaded onto the lanes 2 and 4 as indicated.

The protein entry names of the targets (left) and their corresponding molecular weight (right) are listed.

IHC image of beta Actin staining in human normal colon formalin fixed paraffin embedded tissue section*.

The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH 6) for 30 mins. The section was incubated with ab13822, 10 µg/ml overnight at +4°C. An HRP-conjugated secondary (ab6877, 1/500 dilution) was used for 1 hr at room temperature. The section was counterstained with hematoxylin and mounted with DPX.

The inset negative control image is taken from an identical assay without primary antibody.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

ab6877 was used at dilution 1/1000 with the primary antibody ab25912 in WB. See the review on ab25912.

ab6877 was used at dilution 1/200 with the primary antibody ab17517 in IHC-P. See the review on ab17517.