SARS-CoV-2 S Protein-Specific IFN-γ ELISpot

PBMCs were isolated from ethylene diamine tetraacetic acid (EDTA) whole blood using LeucosepTM tubes (Greiner Bio-one International GmbH, Frickenhausen, Germany) and Histopaque^®-1077 density gradient cell separation medium (Sigma-Aldrich, St Louis, MO, USA) according to the manufacturers’ instructions. IFN-γ ELISpot assay of PBMCs was performed using the ImmunoSpot^® Human IFN-γ Single-Color Enzymatic ELISpot Assay Kit according to the manufacturer’s protocol (Cellular Technology Limited, Shaker Heights, OH, USA). PBMCs were plated at a concentration of 300,000 cells per well and were stimulated with two contiguous peptide pools spanning the length of the SARS-CoV-2 S protein sequence at a concentration of 2 µg/mL per peptide (Mimotopes, Mulgrave, Australia). Imaging was performed using the CTL ImmunoSpot^® Software (Cellular Technology Limited, Shaker Heights, OH, USA). Spot forming units (SFU) were hand counted and calculated per 10 [6 (link)] PBMCs as summed across the peptide pools for each animal.

Free full text: Click here

van Doremalen N., Fischer R.J., Schulz J.E., Holbrook M.G., Smith B.J., Lovaglio J., Petsch B, & Munster V.J. (2021). Immunogenicity of Low-Dose Prime-Boost Vaccination of mRNA Vaccine CV07050101 in Non-Human Primates. Viruses, 13(8), 1645.

Publication 2021

LeucosepTM tubes Histopaque®-1077 density gradient cell separation medium ImmunoSpot® Human IFN-γ Single-Color Enzymatic ELISpot Assay Kit peptide pools CTL ImmunoSpot® Software

Animal Assay Blood Cell separation Cellular Elispot Enzymatic assay Ethylene diamine tetraacetic acid Histopaque Human Ifn γ Peptide Sars cov 2 s protein

Corresponding Organization : National Institutes of Health

Other organizations : CureVac (Germany)

Top 5 similar protocols

Variable analysis

independent variables

Stimulation of PBMCs with two contiguous peptide pools spanning the length of the SARS-CoV-2 S protein sequence at a concentration of 2 µg/mL per peptide

dependent variables

Spot forming units (SFU) per 10^6 PBMCs as summed across the peptide pools for each animal

control variables

PBMC isolation from EDTA whole blood using Leucosep tubes and Histopaque-1077 density gradient cell separation medium
IFN-γ ELISpot assay of PBMCs using the ImmunoSpot Human IFN-γ Single-Color Enzymatic ELISpot Assay Kit
PBMC plating concentration of 300,000 cells per well
Imaging performed using the CTL ImmunoSpot Software

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

Based on most similar protocols

Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.

As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!