Anti fn1

For immunohistochemical staining, paraffinized sections (5 μm) of PDLSC or JBMSC sheets were deparaffinized; blocked; and incubated for 1 h in the following primary antibodies: (1) anti-FN1 (1:200), (2) anti-COL I (1:200), and (3) anti-TNALP (1:200) (1:200, all from Santa Cruz Biotechnologies, Dallas, TX, USA). PBS was used in place of the primary antibodies for negative controls. The sections were then incubated for 45 min in biotinylated secondary antibodies (1:1000) purchased from ZSGB BIO (Peking, China). Each experiment was repeated in triplicate.

The prepared protein samples (10 μL) from 15 AHT-HIV/AIDS patients, 15 YDSK-HIV/AIDS patients and 15 healthy individuals were treated with gel-loading buffer for SDS-PAGE electrophoresis at 120 V for 1 h. The samples were then transferred to a PVDF membrane (Amersham Biosciences) at 15 V for 20 min. The membranes were blocked with 5% bovine serum albumin at 37 °C for 2 h and then incubated with primary antibodies (including anti-FN1, anti-GPX3, anti-KRT10, anti-RBP4, anti-ApoE and anti-KNG1, 1:500, Santa Cruz Biotechnology, CA, USA) at 4 °C overnight, followed by incubation with alkaline phosphatase-conjugated secondary antibodies (1:1000) at 37 °C for 1 h in darkness. The bands were visualized with enhanced chemiluminescence (GE Healthcare) and the band intensities were quantified with ImageJ (Wayne Rasband, National Institutes of Health). Albumin was used as reference to calculate the relative intensity of each protein. The calculation of mean ± standard deviation and Students’ t tests were performed with GraphPad Prism 6.0 software.

Western blot analyses were performed as previously described [7 (link)]. List of antibodies used: anti-LY75 (Abcam, Branford, CT, USA and Santa Cruz Biotechnology Dallas, TX, USA), anti-Snail1, anti-FN1, anti-E-cad, anti-EpCAM, anti-AXIN1, anti-WNT3, anti-EVX2, anti-IFFO1, anti-CLDN5, anti-HOOK1, anti-JMJD8, anti-RAMP2, anti-KLF-4, anti-β-actin antibodies (Santa Cruz Biotechnology, Dallas, TX, USA) and anti-Twist1, anti-E-cadherin, anti-N-cadherin, anti-APC2, and anti-Cend1 antibodies (Abcam Branford, CT, USA).