Variable expression of cystatin C in cultured trans-differentiating rat hepatic stellate cells

Figure 2 Confocal laser scanning microscopy and ELF assay for quantitative determination of CysC expression in HSCs and MFBs. A: HSCs/MFBs were seeded on glass cover slides for indicated time intervals. After fixation, cells were permeabilized and incubated with primary antibodies directed against CysC or α-SMA. Primary antibodies were visualized with a FITC-labeled secondary antibody (CysC, green fluorescence) or a Cy3-labeled secondary antibody (α-SMA, red fluorescence). Original magnification was ×400. Negative controls, using normal IgGs instead of primary antibodies showed no staining (data not shown); B: HSCs/MFBs cultured in 96-well plates for indicated time intervals were fixed and stained for CysC. The staining was visualized by fluorescence microscopy (original magnification ×200). Fluorescence intensities were measured quantitatively using a 96-well fluorescence reader (excitation 365 nm, emission ≥515 nm).