Fig. 5.

Binding of Hsp70 protein to PDT-treated tumor cells. LLC cells were exposed to Photofrin™ at 10 μg/ml (for PDT1), 20 μg/ml (for PDT2, same dose as in Fig. 1), or 40 μg/ml (for PDT3) for 24 h and then treated with 1 J/cm² of 630 ± 10 nm light. The cells were thereafter incubated in full growth medium at 37°C. At 2.5 h post-PDT light treatment, Hsp70-FITC or albumin-FITC was added at 10 μg/sample and the cells were further incubated for 30 min at 37°C (except for one group incubated at 4°C) before they were collected for flow cytometry analysis. a Hsp70- or albumin-associated fluorescence of cells treated with PDT based on three different Photofrin™ doses and controls including untreated, Photofrin™-only (20 μg/ml), and light-only (1 J/cm²)-treated cells (the fluorescence values were corrected by subtracting the background measured with cells receiving the same treatment but no FITC-labeled protein)—the insert depicts FITC fluorescence-positive fraction size for untreated and PDT2-treated cells exposed to Hsp70-FITC; b Hsp70-associated fluorescence on cells treated with PDT2 identified as viable, apoptotic, or necrotic; c PE fluorescence obtained with annexin V–PE staining of PDT2-treated cells exposed or not exposed to Hsp70-FITC. N = 4, bars are SD; *p < 0.05, statistically significant difference compared to PDT-untreated controls; **p < 0.05, statistically significant difference compared to viable cells in the PDT-treated group; ***p < 0.05, statistically significant difference compared to Hsp70 not added group