Meso Scale Discovery (MSD) kits were used to measure the levels of Aβ42, p-Tau, and t-tau in the RIPA fraction of the human brain. Serial dilutions of the RIPA fractions were tested to measure protein levels accurately. Before performing the assay, protein levels were measured using a Thermofisher BCA Assay Kit. With those results, 1 μg of protein from the soluble fraction was diluted to evaluate Aβ42, t-tau and p-Tau levels. The plates were developed and read using a QuickPlex Q120 reader (Meso Scale Diagnostics). ELISA plates from Abcam (ab269555) were used to measure the levels of Gal-3 (detection range 58.8–2000 pg/ml) in tissue homogenates. The protocol was carried out according to the manufacturer’s instructions. A Biotek Synergy 2 was used to read the ELISA Gal-3 assay.
Ab269555
Manufactured by Abcam
Ab269555 is a lab equipment product offered by Abcam. It is a device designed for use in scientific research applications. The core function of this product is to perform a specific task or activity within a laboratory setting. No further details about the intended use or features of this product can be provided in an unbiased and factual manner.
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3 protocols using ab269555
1
Quantifying Alzheimer's Biomarkers and Gal-3 in Human Brains
2
ELISA for Galectin-3 Quantification
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Enzyme‐linked immunosorbent assay (ELISA) plates from Abcam (ab269555) were used to measure the levels of Gal‐3 (detection range 58.8 to 2000 pg/mL) in tissue homogenates, CSF, and serum samples. The protocol was carried out according to the manufacturer's instructions. A Biotek Synergy 2 was used to read the ELISA Gal‐3 assay. All samples were run in duplicate once. Mean inter‐assay Coeficients of Variances (CV) was 6.23. Samples were distributed in the plates according to the clinical group in similar proportions to avoid a bias caused by the plate. The ELISA and the analysis of the raw data were performed by different persons. CSF samples were not diluted. Tissue homogenates were diluted 1:100. Serum samples were diluted 1:20 or 1:50; the correction factor on diluted samples was applied when needed for the comparison. Our kit has been used previously to measure Gal‐3 levels in brain, CSF, and serum samples
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3
Quantification of Secreted Galectins
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Cell suspensions were centrifuged at 300× g for 5 min and the supernatants (8–10 mL) were collected and frozen at −80 °C to use later for analyses of extracellular proteins. Concentrations of secreted galectin-1, galectin-3, and galectin-9 were measured using ELISA kits from Abcam (ab260053, ab269555, and ab213786) as described by the manufacturer. For all three kits, cell culture supernatant samples were diluted 4-fold in the supplied sample diluent that was found to fall within the range of concentrations that correspond to the standard curve. Absorbance at 450 nm was measured using a Model 3550 Microplate Reader (Bio-Rad).
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