Fig 1: Dynamic range of CA9 detection using LFA strips. (A) Nine test solutions were prepared by spiking recombinant CA9 into benign seroma samples at defined concentrations, using a 1:2 ratio of seroma to running buffer. Results were imaged using both an iPhone camera (color) and a Bio-Rad imager (black and white) and image analysis was performed using NIH Image J software. (B) The TL/CL ratio exhibited a strong linear correlation with CA9 concentration (Y = 0.13X + 0.45, r2 = 0.96, p < 0.0001). CL, control line; TL (CA9), test line.
Fig 2: CA9 Expression in TLBR-1 and TLBR-2 Cell Lines. (A) Immunofluorescence staining of TLBR-1 and TLBR-2 cell lines showed CA9 expression (green), with nuclei counterstained with DAPI in blue. Goat IgG was used as a negative control. (B) LFA analysis demonstrated the presence of CA9 in culture supernatants of both cell lines. CL, control, line; Scale bar = 100 μm.
Fig 3: Heatmap comparing concentrations of CA9, CD30 and 12 cytokines in BIA-ALCL and benign seromas.
Fig 4: Statistical analysis of CA9 LFA performance. (A) A strong linear correlation was observed between CA9 concentrations measured by ELISA and the TL/CL ratio from the LFA (Y = 0.18X − 0.12, r2 = 0.86, p < 0.0001). (B) CA9 levels were significantly elevated in BIA-ALCL samples compared to benign seromas (Benign: 0.38 ± 0.06, n = 23; BIA-ALCL: 0.67 ± 0.19, n = 28; t-test, **** p < 0.0001). (C) Receiver Operating Characteristic (ROC) curve analysis yielded an area under the curve (AUC) of 0.95, indicating excellent diagnostic performance.
Fig 5: CA9 LFA analysis on benign and BIA-ALCL seromas. Images were captured using an iPhone camera (color) and the Bio-Rad imager with the Alexa Fluor 488 channel (black and white). Density of CL and TL were determined from Bio-Rad images.
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