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Clone 2.4g2

Manufactured by BioXCell
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The Clone 2.4G2 is a laboratory product designed for use in cell biology research. It functions as an antibody that binds to the mouse Fc gamma receptor II/III (CD16/CD32). The core purpose of this product is to facilitate the study of immune cell interactions and signaling pathways.

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Lab products found in correlation

Rbc lysis buffer, by Thermo Fisher Scientific (2 mentions) Hank s balanced salt solution, by Thermo Fisher Scientific (2 mentions) 100 and 40 mm nylon cell strainers, by BD (2 mentions) 70 and 40 mm nylon cell strainers, by BD (2 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Dnase 1, by Roche (2 mentions) Fitc conjugated anti b220, by BD (1 mentions) Alexa fluor 647 fluorochrome, by Thermo Fisher Scientific (1 mentions) Alexa fluor 750, by Thermo Fisher Scientific (1 mentions) Facscanto, by BD (1 mentions) Rat igg2a isotype control clone 2a3, by BioXCell (1 mentions)

4 protocols using clone 2.4g2

1

Epitope Mapping of Conjugated Antibodies

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Purified hcAbs were conjugated via amino groups to Alexa Fluor647-fluorochrome according to the manufacturer’s instructions (Molecular Probes, Thermo Fisher Scientific). For epitope mapping analyses, EL4 cells were pre-incubated with a saturating concentration (100 nM) of unconjugated hcAbs for 30 min at 4°C, followed by addition of Alexa Fluor647-conjugated hcAbs (10 nM) and further incubation for 20 min at 4°C. Cells were washed and analyzed by flow cytometry on a BD-FACS Canto. Data was analyzed using the FlowJo software (Treestar). The percentage of cross-blockade was calculated from mean fluorescence intensities (MFI) as follows: (MFI in the absence of competing Abs – MFI in the presence of competing Abs): (MFI in the presence of competing Abs) x 100. Spleen cells were pre-incubated with Fc-block (BioXcell, clone 2.4G2) to minimize unspecific binding to Fc-receptors. Cells were then incubated with Alexa Fluor647-conjugated hcAbs, FITC-conjugated anti-B220 (BD biosciences, clone RA3-6B2), and Alexa Fluor 750 as a viability dye (ThermoFischer). Gating was performed on Alexa Fluor 750-low cells (live cells).
Baum N., Eggers M., Koenigsdorf J., Menzel S., Hambach J., Staehler T., Fliegert R., Kulow F., Adam G., Haag F., Bannas P, & Koch-Nolte F. (2021). Mouse CD38-Specific Heavy Chain Antibodies Inhibit CD38 GDPR-Cyclase Activity and Mediate Cytotoxicity Against Tumor Cells. Frontiers in Immunology, 12, 703574.
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2

Isolation of Murine and Human PDAC Cells

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Mouse and human PDAC tumors and adjacent pancreata were mechanically dissociated and incubated in collagenase (collagenase II for murine tumors, collagenase IV for human tumors, both 5 mg/ml; Worthington Biochemical Corp., Fisher Scientific), DNAse I (0.5 mg/ml; Roche Diagnostics), and Hank’s balanced salt solution (Gibco, Fisher Scientific) for 30 minutes at 37°C. Digestion was then quenched with fetal bovine serum (FBS, Life Technologies), and cells were filtered sequentially through 100- and 40-mm nylon cell strainers (Falcon, Fisher Scientific). Tumors, adjacent pancreata, and lymph nodes were then mechanically disassociated and filtered through 100- and 40-mm nylon cell strainers (Falcon, Fisher Scientific) using PBS with 1% FBS (Life Technologies). Spleens were mechanically dissociated and filtered through 70- and 40-mm nylon cell strainers (Falcon, Fisher Scientific) using PBS with 1% FBS, followed by RBC lysis (RBC lysis buffer, ThermoFisher Scientific). Mouse Fc receptors were blocked with FcεRIII/II-specific antibody (1 μg per 1 × 106 cells; clone 2.4G2, Bio X Cell).
Moral J.A., Leung J., Rojas L.A., Ruan J., Zhao J., Sethna Z., Ramnarain A., Gasmi B., Gururajan M., Redmond D., Askan G., Bhanot U., Elyada E., Park Y., Tuveson D.A., Gönen M., Leach S.D., Wolchok J.D., DeMatteo R.P., Merghoub T, & Balachandran V.P. (2020). ILC2s amplify PD-1 blockade by activating tissue-specific cancer immunity. Nature, 579(7797), 130-135.
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3

Murine Tumor Growth Monitoring and Interventions

Cited 1 time
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Animal research was performed in accordance with the Institutional Animal Care and Use Committees at MGH. DPE-GFP mice (21 (link)) were generously provided by Ulrich von Andrian (Dept. of Microbiology and Immunobiology, Harvard Medical School), IFN-γ reporter with endogenous polyA tail (GREAT) mice (22 (link)) were kindly provided by Andrew Luster (Division of Rheumatology, Allergy and Immunology, Massachusetts General Hospital). Tumor growth was monitored by caliper measurement and the area (A) of these predominantly two-dimensional tumors was calculated using the formula A = length * width. Tumor implantation was performed by intradermal injection of tumor cells (2 × 106 MC38/MC38-H2B-mApple, 5 × 105 B16-F1-Ova, and 5 × 105 KP1.9). MC38 cells were gifted by Dr. Mark Smyth (QIMR Berghofer, Brisbane, Australia), B16 cells were from ATCC, and KP1.9 cells were a gift from Dr. Alfred Zippelius (University Hospital Basel, Switzerland). Experiments were generally started when tumors became vascularized, which was after 8 days. For aPD-1 and AF647-aPD-1 treatments, mice were given 200 μg i.p. of the 29F.1A12 aPD-1 clone. For in vivo Fc blocking experiments, mice were infused i.p. with 200 μg of monoclonal antibody specific to mouse Fc gamma receptors II and III (clone 2.4G2, Bioxcell) daily for 5 days. Control mice received 200 μg rat IgG2a isotype control (clone 2A3, Bioxcell).
Arlauckas S.P., Garris C.S., Kohler R.H., Kitaoka M., Cuccarese M.F., Yang K.S., Miller M.A., Carlson J.C., Freeman G.J., Anthony R.M., Weissleder R, & Pittet M.J. (2017). In vivo imaging reveals a tumor-associated macrophage mediated resistance pathway in anti-PD-1 therapy. Science translational medicine, 9(389), eaal3604.
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4

Isolation of Murine and Human PDAC Cells

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Mouse and human PDAC tumors and adjacent pancreata were mechanically dissociated and incubated in collagenase (collagenase II for murine tumors, collagenase IV for human tumors, both 5 mg/ml; Worthington Biochemical Corp., Fisher Scientific), DNAse I (0.5 mg/ml; Roche Diagnostics), and Hank’s balanced salt solution (Gibco, Fisher Scientific) for 30 minutes at 37°C. Digestion was then quenched with fetal bovine serum (FBS, Life Technologies), and cells were filtered sequentially through 100- and 40-mm nylon cell strainers (Falcon, Fisher Scientific). Tumors, adjacent pancreata, and lymph nodes were then mechanically disassociated and filtered through 100- and 40-mm nylon cell strainers (Falcon, Fisher Scientific) using PBS with 1% FBS (Life Technologies). Spleens were mechanically dissociated and filtered through 70- and 40-mm nylon cell strainers (Falcon, Fisher Scientific) using PBS with 1% FBS, followed by RBC lysis (RBC lysis buffer, ThermoFisher Scientific). Mouse Fc receptors were blocked with FcεRIII/II-specific antibody (1 μg per 1 × 106 cells; clone 2.4G2, Bio X Cell).
Moral J.A., Leung J., Rojas L.A., Ruan J., Zhao J., Sethna Z., Ramnarain A., Gasmi B., Gururajan M., Redmond D., Askan G., Bhanot U., Elyada E., Park Y., Tuveson D.A., Gönen M., Leach S.D., Wolchok J.D., DeMatteo R.P., Merghoub T, & Balachandran V.P. (2020). ILC2s amplify PD-1 blockade by activating tissue-specific cancer immunity. Nature, 579(7797), 130-135.
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