Journal: Methods in cell biology

Article Title: Standardized protocol for the evaluation of chimeric antigen receptor (CAR)-modified cell immunological synapse quality using the glass-supported planar lipid bilayer

doi: 10.1016/bs.mcb.2022.07.009

Figure Lengend Snippet: d

Article Snippet: Reagent or resource Source Identifier Antibodies Alexa-Fluor 405 Phalloidin Thermofisher A30104 Alexa-Fluor 568 mouse anti-perforin (dG9) Biolegend 308102 Alexa Fluor 647 rabbit anti-CD3 zeta (phosphor Y83) Abcam Ab68236 Chemicals, Peptides, and Recombinant Proteins 18:1(Δ-Cis) PC (DOPC) 1,2-dioleoyl-snglycero-3-phosphocholine (10mg/mL) Avanti Polar Lipids 850375C 18:1 Biotinyl Cap PE 1,2-dioleoylsnglycero-3-phosphoethanolamine-N-(cap biotinyl) (10mg/mL) Avanti Polar Lipids 870273C Compressed Argon gas Airgas AR UHP180LT230 Sulfuric Acid Acros Organics 7664-93-9 Hydrogen peroxide Sigma 386,90 Tris Fisher Scientific BP152-1 Potassium hydroxide Sigma P5405 Sodium chloride Fisher Scientific S5886-10KG n-octyl-β- -glucopyranoside Sigma O8001 HEPES Sigma H4034 30% human serum albumin (HSA) Sigma 12667-50ML-M Streptavidin Thermofisher 434,301 IgG1, Kappa from human myeloma plasma Sigma I5154-1MG D-Biotin Thermofisher B20656 32% paraformaldehyde VWR 100504-858 Normal donkey serum (NDS) Jackson Immunoresearch Laboratories 017-000-121 Triton-X Sigma 9036-19-5 190 Proof ethanol Fisher Scientific 64-17-5 10× PBS Fisher Scientific MT-46013CM Storage glassware equipment Sample vials, Boroscilicate glass VWR 14230-826 Experimental Models: Cell Lines Kappa CAR NK92MI cells Generated in-house Equipment Lyophilizer Labconco Freezone 7740020 Sonicator QSonica Q500 Dialysis tubing Fisher Scientific 08-670-5C Vapro Osmometer ELITechGroup Model 5600 IBIDI Sticky-side VI 0.4 6-well chambers IBIDI 80608 25x75mm glass coverslips IBIDI 10812 Nikon A1R HD Confocal microscope Nikon Software and Algorithms Image J (Fiji) NIH Graphad Prism 9.3.1 Graphpad Other Machine learning-based CAR IS quality analysis Developed in-house See PLOS Computational Biology ( Naghizadeh et al., 2022 ).

Techniques: Recombinant, Clinical Proteomics, Generated, Microscopy, Software