d) Intracellular IgG1 staining with and without blocking surface IgG1 with an unlabeled antibody. Supplementary information Supplementary Information Supplementary Tables 1–3, Text and Methods. Reporting Summary Supplementary Software Programmed Excel worksheet that performs calculations for this protocol. ...
The development of MHC class I and II tetramer staining techniques allows precise quantification and tracking of antigen-specific CD8(+) and CD4(+) T cell responses. Here, we describe a protocol for MHC class I and II tetramer staining of mouse T cells isolated from various tissues of mice ...
IHC staining of resected mouse tumor tissues and human lung cancer tissue arrays was carried out according to the manufacturer’s protocol (Zhongshan Golden Bridge, Beijing, China). The quantification of IHC was calculated as IRS (0–12) = SP (0–1: 0%–25%; 2: 26%–50%; 3: 51...
Lastly, the native gel was silver stained using SilverQuest Silver Staining Kit (Invitrogen) according to the manufacturer’s protocol. Formaldehyde and BS3 Crosslinking TRPM4-eGFP was crosslinked using the imidoester crosslinkers formaldehyde (FA, Sigma-Aldrich) and bis-sulfosuccinimidyl suberate (...
Peptide exchange Basic Protocol 5: Analysis of peptide exchange by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry Alternate Protocol 2: Native isoelectric focusing to validate MHC-II peptide loading Basic Protocol 6: Multimerization Basic Protocol 7: Staining cells with Class II ...
Additionally, these MHC-II/peptide complexes that interact with T-cell receptors (TCR) of pathogenic CD4+ T cells are compatible with a different number of assays like Intracelullar Cytokine Staining (ICS), and Carboxyfluorescein succinimidyl ester (CFSE) making it a robust technology to study ...
Purified proteins were resolved on 7.5–20% sodium dodecyl sulphate-polyacrylamide gel electrophoresis gradient gels (Wako) and stained with silver staining. Electrophysiology of mammalian cells. All electrophysiological experiments were performed as described previously7. The pipette solution contained 105 ...
at the beginning of metaphase, peroxisomes began to congregate in two clusters on the bottom and top of the emerging metaphase plate (FIG. 2D). Additional immunochemical staining of the mitotic spindle revealed that the peroxisomes
Identification of Human Antigen-Specific T Cells Using Class II MHC Tetramer Staining and EnrichmentAntigen-specific T cellsCD4+ T cellsHuman immune responseImmune repertoireMHC class IIT cell enrichmentTetramersThe development of peptide-major histocompatibility complex tetramers has enabled direct ...
The optimal procedure for assessing the functional activity of tetramer-binding CD4+ T cells was based on the simultaneous intracellular staining with both MHC tetramers and cytokine-specific antibodies upon in vitro restimulation of cells with the vaccine antigen. The protocol was selected among ...