ELISPOT assays were performed according to previously described methods [13 (link),15 (link)]. To quantitate influenza-specific antibody secreting cells (ASC), the multiscreen 96-well HTS plates (Millipore) were coated with TIV at 5μg/ml HA protein of each strain. Thawed PBMCs were re-suspended in complete RPMI 1640 culture medium, counted and plated at the density of 0.2×106 cells per well and cultured overnight. The plates were then washed and probed with anti-human IgG-HRP, IgM-HRP or IgA-HRP. After a final wash the spots were developed with AEC Substrate set (BD Biosciences) and enumerated on an ELISPOT reader (Cellular Technology Ltd). The baseline background was subtracted to obtain the 7dpv ASC frequency.
Hts plates
Manufactured by Merck Group
HTS plates are multi-well microplates used in high-throughput screening (HTS) applications to facilitate the rapid and automated testing of large numbers of chemical compounds or biological samples. These plates are designed to provide a standardized format for conducting parallel assays, enabling efficient data collection and analysis.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using hts plates
1
Quantifying Influenza-Specific Antibody Secreting Cells
2
LIPS Assay for COVID-19 Antibodies
Check if the same lab product or an alternative is used in the 5 most similar protocols
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The LIPS assays were performed following the protocol of Burbelo et al., with the following modifications36 (link), as previously described14 (link). Briefly, (Ruc)-antigen (at an equal concentration for each antigen at 10^7 per well) and plasma (heat inactivated and diluted 1:100) were incubated for 2 hours with shaking at 800rpm. Ultralink protein A/G beads (Thermo-Fisher) were added to the (Ruc)-antigen and serum mixture in a 96-deep-well polypropylene microtiter plate and incubated for 2 hours with shaking at 800rpm. The entire volume was then transferred into HTS plates (Millipore) and washed as previously described. The plate was read using QUANTI-Luc Gold substrate (Invivogen) as per manufacturer’s instructions on a Wallac MicroBeta JET luminometer 1450 LSC & Luminescence counter and its software for analysis (PerkinElmer). Experimental controls include no plasma blank wells with (Ruc)-antigens and negative control serum from healthy donors plasma collected prior to the COVID-19 pandemic. The background corresponds to the LU signal from each Ruc-fusion antigen with protein A/G and substrate with no plasma.
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