Secreted tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β), TNF-α, IL-6, and IL-1β concentrations in cultured supernatants were analyzed at 6 h OGD and 12 h OGD and reperfusion (OGD/R) via ELISA (DuoSet Mouse, R&D Systems) in 96-well plates prepared according to manufacturer's instructions. Briefly, captured antibody-precoated plates were incubated with blocking solution for 1 h at room temperature, supernatants were collected and transferred in duplicates onto a 96-well-plate and incubated overnight at 4°C, and detection antibodies were transferred at a dilution 1 : 60 and incubated for 3 h, followed by light-protected incubation with HRP-conjugated streptavidin for 20 minutes at room temperature. Extinction was measured at 450 nm and 540 nm using a microtiter plate reader (Thermo Fisher Multiskan Ascent).
Duoset mouse
Manufactured by R&D Systems
Sourced in United States
The DuoSet Mouse is a laboratory equipment product designed for sandwich ELISA assays. It contains the necessary components, including capture and detection antibodies, to measure the concentration of specific mouse proteins in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using duoset mouse
1
Cytokine Quantification in OGD/R
2
Retinal Protein Isolation and IL-1β Quantification
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Retinas were collected as described previously.38 (link) Equal amounts of protein from cell lysates or retinal homogenates were combined with Laemmli buffer, boiled for 5 min, and fractionated using a 4-20% Criterion TGX Precast gel (Bio-Rad Laboratories). Proteins were transferred to a polyvinylidene fluorine membrane, blocked using 5% milk in Tris-buffered saline Tween 20, and incubated with antibodies (Supplementary Table S1 ). The concentration of IL-1β was determined in the retina whole cell lysates or in culture medium using the DuoSet mouse (DY401-05) and human (DY201-05) IL-1β/IL-1F2 ELISA kits (R&D Systems, Minneapolis, MN, USA), respectively.
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