Journal: bioRxiv

Article Title: Cellular thermal shift assay of subcellular isolates for evaluating drug-membrane target interactions

doi: 10.64898/2026.02.03.703651

Figure Lengend Snippet: (A) Western blot image of MPC1 at 12 kD in the soluble fraction from 37 to 66°C of whole cells treated with vehicle control and UK5099. (B) The western blot was quantified using Image Lab.Relative intensities were used to quantify protein abundance normalized to 37°C. Curves fitted to the Boltzmann Sigmoid equation and demonstrate no statistically significant difference between the melting temperature of MPC1 in the control and UK5099-treated conditions (n = 1, N = 3, extra sum of squares F test, p = 0.91). (C) Western blot image of MPC at 12 kD in the soluble fraction of whole cell lysate treated with vehicle control and UK5099. (D) Quantified protein abundance in the soluble fraction fitted to the Boltzmann Sigmoid equation demonstrate no significant difference in the melting temperatures between control and UK5099-treated conditions (n = 1, N = 3, extra sum of squares F test, p = 0.82). Blot images are representative of means obtained from independent replicates.

Article Snippet: The membrane was blocked in 2% BSA for two days before being probed with 1:500 MPC1 antibody (Cell Signaling Ref 14462) [ ], 1:2000 MPC2 antibody (Cell Signaling Ref 46141) [ ], 1:2000 Porin VDAC antibody (Calbio Ref 529534), or 1:2000 ß-actin antibody (Cell Signaling Ref 3700) overnight.

Techniques: Western Blot, Control, Quantitative Proteomics

Journal: bioRxiv

Article Title: Cellular thermal shift assay of subcellular isolates for evaluating drug-membrane target interactions

doi: 10.64898/2026.02.03.703651

Figure Lengend Snippet: (A) Western blot image of MPC1 in the soluble fraction between the temperatures of 37 and 66°C in DMSO and UK5099-treated conditions. (B) The western blot was quantified as described above. The DMSO samples were fitted to a Boltzmann Sigmoid curve and compared to the UK5099 samples (n = 1-2, N = 3, paired t-test assuming unequal variance, * = p < 0.05, ** = p < 0.01). (C) Western blot image of MPC2 in the soluble fraction between the temperatures of 37 and 66°C in DMSO and UK5099-treated conditions. (D) The western blots were quantified as above and the DMSO samples were fitted to a Boltzmann Sigmoid curve and compared to the UK5099 samples (n = 1-2, N = 3, extra sum of squares F-test, p < 0.05). Blot images are representative of means obtained from independent replicates.

Article Snippet: The membrane was blocked in 2% BSA for two days before being probed with 1:500 MPC1 antibody (Cell Signaling Ref 14462) [ ], 1:2000 MPC2 antibody (Cell Signaling Ref 46141) [ ], 1:2000 Porin VDAC antibody (Calbio Ref 529534), or 1:2000 ß-actin antibody (Cell Signaling Ref 3700) overnight.

Techniques: Western Blot

Journal: bioRxiv

Article Title: Cellular thermal shift assay of subcellular isolates for evaluating drug-membrane target interactions

doi: 10.64898/2026.02.03.703651

Figure Lengend Snippet: (A) Schematic representing hypothesis that cellular lysis disrupts mitochondrial membrane integrity whereas with use of mitochondrial isolation it is preserved. (B) Western blot image of MPC1 in the soluble fraction between the temperatures 37 and 66°C in control versus UK5099 treated samples. (C) The samples were fitted to the Boltzmann Sigmoid curve, and no significant difference was observed in the MPC1 melting temperature between control and UK5099-treated conditions (n = 1-2, N = 3, extra sum of square F test, p = 0.7611). Blot images are representative of means obtained from independent replicates.

Article Snippet: The membrane was blocked in 2% BSA for two days before being probed with 1:500 MPC1 antibody (Cell Signaling Ref 14462) [ ], 1:2000 MPC2 antibody (Cell Signaling Ref 46141) [ ], 1:2000 Porin VDAC antibody (Calbio Ref 529534), or 1:2000 ß-actin antibody (Cell Signaling Ref 3700) overnight.

Techniques: Lysis, Membrane, Isolation, Western Blot, Control