For confocal microscopy, conjugates between CAR T cells and target cells were incubated for 30 minutes at 37°C and then fixed and stained for F-actin (Phalloidin 568), Pericentrin (rabbit anti-Pericentrin; Abcam), and ICAM-1 (mouse anti-ICAM-1; BD Biosciences). Pericentrin was detected using Alexa Fluor Pacific Blue secondary antibody to rabbit IgG, and ICAM-1 was detected using 488 Alexa Fluor secondary antibody to mouse IgG. Conjugates were imaged as Z stacks of 0.2 μm thickness to cover the entire volume of the immune synapse (IS), determined individually for each conjugate on a Leica SP8 confocal microscope using a 100× objective. Density of F-actin and ICAM-1 accumulation at the immune synapse of CAR T cells was calculated using the formula [volume × mean fluorescence intensity (MFI)] for an equal number of 1 μm3 boxes selected to cover the IS (19 (link)). Microtubule-organizing center (MTOC) to synapse distance was measured as described before (20 (link)).
Mouse anti icam 1
Manufactured by BD
Mouse anti-ICAM-1 is a laboratory reagent used for the detection and quantification of the Intercellular Adhesion Molecule 1 (ICAM-1) in research samples. ICAM-1 is a cell surface glycoprotein involved in the adhesion of cells to other cells.
Automatically generated - may contain errors
Lab products found in correlation
Pericentrin, by Abcam (1 mentions)
Anti pericentrin, by Abcam (1 mentions)
Sp8 confocal microscope, by Leica (1 mentions)
Ve cadherin, by Santa Cruz Biotechnology (1 mentions)
Anti e selectin, by BD (1 mentions)
Goat anti claudin 5, by Santa Cruz Biotechnology (1 mentions)
Rabbit anti gapdh, by Abcam (1 mentions)
Anti vcam 1, by BD (1 mentions)
2 protocols using mouse anti icam 1
1
Visualizing Immune Synapse Formation
2
Immunoassay Antibody Characterization Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Mouse anti-HRPII (2G12), a generous gift from Diane Taylor (University of Hawaii), was used at 1:100, goat anti-claudin-5 (from Santa Cruz; sc-17667) at 1:100, and VE-cadherin (Santa Cruz; sc-52751) at 1:100. Five micrograms of anti-HRPII was used for neutralization and was from Thermo Scientific (MA1-27094), and the isotype control (269) was generated as described previously (65 (link)). Rabbit anti-IL-1β (Rockland Immunochemical; 209-401-301) was used for neutralization. Rabbit anti-GAPDH (anti-glyceraldehyde-3-phosphate dehydrogenase) (Abcam; ab37168) was used at 1:1,000. Mouse anti-ICAM-1, anti-VCAM-1, and anti-E-selectin (BD Biosciences; 555510, 555645, and 555648) were used at 1:500, 1:100, and 1:500 dilutions, respectively. Armenian hamster anti-IL-1β (Leinco) was used for in vivo neutralization at 300 µg/mouse.
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