tnf α production assay  (Elabscience Biotechnology)

Journal: mAbs

Article Title: Higher order receptor clustering due to the IgG3 subclass is necessary for TLR4 signaling and tolerance induction by novel human anti-TLR4 antibodies

doi: 10.1080/19420862.2025.2515415

Figure Lengend Snippet: Novel human IgG4 antibodies 3D7, 8G10, and 8H1 recognize TLR4 and MD2 while 7D11 recognizes only MD2. Detection of recombinant TLR4/MD2 heterodimer (a) TLR4 alone (b) or MD2 alone (c) by ELISA using a 3-fold dilution series of human IgG4 antibodies 3D7, 8G10, 8H1, and 7D11. With the concentration of TLR4Ab held constant, an increasing concentration of soluble TLR4 was added in the ELISA assay to compete with the plate bound TLR4/MD2 (d). Data represents a single experiment in duplicate.

Article Snippet: Supernatants were again collected at 24 h and assayed for TNF-α production by ELISA (R&D Systems, #DY210–05).

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: mAbs

Article Title: Higher order receptor clustering due to the IgG3 subclass is necessary for TLR4 signaling and tolerance induction by novel human anti-TLR4 antibodies

doi: 10.1080/19420862.2025.2515415

Figure Lengend Snippet: IgG3 TV-7D11 induces tolerance in primary human myeloid cells. Human CD14+ monocytes were purified by magnetic bead negative selection. 1 × 10^6 cells/well were seeded in 24-well plates and stimulated with IgG3 8H1 (10 μg/ml), IgG3 TV-7D11 (10 μg/ml), or LPS (.01 μg/ml). After 24 h supernatants were removed and tested for TNF-α by ELISA (a), followed by addition of fresh media and culture for 5 d to allow cells to return to a resting state (b). On day 6, cells were split into 96-well plates at 50,000 cells/well and allowed to rest for 24 h prior to re-challenge with LPS at .01 μg/ml. Supernatants were again collected at 24 h and assayed for TNF-α production by ELISA (c). Data points represent three individuals from two independent experiments. Statistical analysis by ANOVA * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Supernatants were again collected at 24 h and assayed for TNF-α production by ELISA (R&D Systems, #DY210–05).

Techniques: Purification, Selection, Enzyme-linked Immunosorbent Assay