At the time of sacrifice, blood was collected by heart puncture into EDTA-containing tubes and centrifuged at 3,000 rpm for 25 mins to isolate the plasma. Plasma was aliquoted and stored frozen at −80°C. The following ELISA plates from Abcam® (Cambridge, MA) were used for analysis: Fibrinogen, Neutrophil Elastase, C-reactive protein and IL-6.
Elisa plates
Manufactured by Abcam
Sourced in United Kingdom, United States
ELISA plates are a type of multi-well laboratory equipment used for enzyme-linked immunosorbent assay (ELISA) experiments. The plates are designed to hold and process multiple samples simultaneously, allowing for efficient and standardized immunoassay testing.
Automatically generated - may contain errors
3 protocols using elisa plates
1
Plasma Biomarker Quantification Protocol
2
Cytokine Profiling by ELISA
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100 μL cell culture supernatants were added into the ELISA plates (Abcam, Cambridge, UK) pre-coated with PEG2, TGF-β, IDO, TSG6, TNF-α, IL-1β, and IL-6 antibodies and incubated for 2 h at room temperature. After washing, biotin-labeled detection antibody was added and incubated for 1 h at 37 °C. Avidin-horseradish peroxidase was used for color development. The OD values at 450 nm were measured using a microplate reader (Thermo Fisher Scientific).
3
SARS-CoV-2 Spike Protein ELISA Assay
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An ELISA assay assessing binding antibody levels against the wild-type SARS-CoV-2 was conducted using the wild-type Spike protein. ELISA plates (Corning Costar; Corning, NY, USA) were coated with S protein (Sanyou Biopharmaceuticals Co., Shanghai, China) at a concentration of 5 μg/well and incubated overnight at 4 °C. During the experiment, ELISA plates were blocked with 5% BSA-phosphate-buffered saline (PBS), incubated with serially diluted serum samples, and visualized by reaction with an HRP-conjugated antibody (Abcam, Waltham, MA, USA) and TMB substrate (Solarbio, Beijing, China), using previously described methods [16 (link)]. The absorbance of each well at 450 nm was measured using an ELISA plate reader (Gene Company, Beijing, China). Antibody serum samples yielding OD values at least 2.1-fold higher than the negative control at a test sample dilution of 1:400 were considered positive. Endpoint titers (ETs) were defined as the highest serum dilutions yielding positive OD values. GMTs were calculated as the geometric mean of the ETs of positive serum samples in each group. ELISA assays assessing binding antibody levels against variant strains of SARS-CoV-2 (Alpha, Beta, Gamma, Delta, and Omicron) were conducted using commercial ELISA kits (ACRO Biosystems Co., Beijing, China) according to the manufacturer’s instructions.
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