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Background When dealing with solid tumors lacking specific biomarkers, the effectiveness of adoptive T cell therapy using TCR-T treatments is limited. We believe that the absence of dependable, multifaceted MHC reagents has been a hindrance in the research and development of treatments for solid tumors. To address this unmet need, we have engineered MHC molecules in several formats with diverse applications and high stability. 1–7 These improved, highly active MHC molecules have the potential to facilitate solid tumor research and development.Methods Form 1: Mammalian-Expressed Single-Chain Trimer (SCT) MHC for High SensitivityLinkers in SCT maintain MHC integrity and antigen presentation.Some concern exists that linkers may interfere with TCR, but structural studies of 200+ complexes show otherwise.SCT MHCs are mammalian-expressed—not refolded from E. coli—offering improved stability and sensitivity.Form 2: Chimeric MHCs for Reduced Off-Targetsα3 and β2m domains substituted with murine equivalents.Maintains antigen specificity and allows for TCR mimic (TCRm) antibody development.Reduced non-specific binding improves screening efficiency.Form 3: Peptide-Ready MHC (prMHC)No pre-loaded peptides—researchers can load any peptide.Saves time, maximizes flexibility.Ideal for SPR, BLI, ELISA, and T-cell sorting.Protocol Summary:Dissolve peptide in DMSO/PBS.Incubate 5–10 min at RT.Ready to use.Form 4: Biotinylated MHC Monomers & Fluorescent MHC TetramersTetramers allow enhanced detection of antigen-specific T cells.Available as pMHC or prMHC, produced in vivo or in vitro.Compatible with FACS analysis for sorting and quantification.Results Case Study HighlightsCase Study 1: W6/32 mAb validates mammalian-expressed prMHCs and SCT MHCs.prMHCs with native N-terminus β2m are recognized.SCT and E. coli refolded MHCs are not recognized.Case Study 2: prMHC shows activity comparable to SCT MHC when loaded with KRAS G12D.Case Study 3: In vivo-formed MHC tetramers are equivalent to standard tetramers in binding performance.Case Study 4: In vivo-formed HLA-G tetramer binds LILRB2 with strong affinity (IC5 0 = 62.7 ng/mL), outperforming in vitro tetramer (IC5 0 = 0.45 µg/mL).Case Study 5: prMHCs retain activity after lyophilization, reconstitution, and freeze-thaw cycles.Conclusions KACTUS prMHCs support common HLA types (e.g., A02:01, A11:01, A*03:01).User-friendly format for customizable assays.Applications include TCR binding studies and T-cell sorting.Outperforms competitors in tetramer performance and stability.References Braud VM, et al. Nature. 1998;391:795–799.Bylinska A, et al. Mol Genet Genomics. 2018;293:601–613.Hansen TH, et al. Trends Immunol. 2010;31:363–369.Li D, et al. PLoS One. 2017;12:e0176642.Poole A, et al. Nat Commun. 2022;13:5333.Vaurs J, et al. Sci Rep. 2021;11:17234.Shields M, et al. Tissue Antigens. 1998;51(5):567–70.