Western blot analysis of CCR7 was performed by loading the indicated amounts of Jurkat whole cell lysate and 10ul of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a Novex® 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane using the G2 Fast Blotter (Product # 62288), and blocked with 5% nonfat dry milk in TBST for at least 1 hour at room temperature. CCR7 was detected at ~43kD using a CCR7 monoclonal antibody (Product # MA1-163) at a dilution of 1:500 in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-rat IgG secondary antibody (Product # 31470) at a dilution of 1:40,000 for at least 30 minutes at room temperature. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34075).
|Tested species reactivity||Human, Mouse|
|Host / Isotype||Rat / IgG2a|
|Immunogen||Rat basophilic leukaemia (RBL)-2H3 transfectants expressing mouse CCR7|
|Storage buffer||PBS with 1mg/ml BSA, 30% glycerol|
|Contains||0.05% sodium azide|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||5ug/1X10^6 cells|
|Western Blot (WB)||1:250-1:1000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
MA1-163 has been validated for flow cytometry on primary mouse splenocytes. Optimal staining is observed when incubating the cells with antibody at 37C for at least 30 minutes.
MA1-163 has been validated for Western blot on Jurkat cell lysates.
CCR7 is a member of the G protein coupled receptor family (subfamily: chemokine). This receptor was identified as a gene induced by the Epstein Barr virus (EBV), and is thought to be a mediator of EBV effects on B lymphocytes. CCR7 has been reported to be expressed in blood, bone marrow, lymph node, and intestine. It is particularly expressed in lymphoid tissues and in activated B and T lymphocytes and has been shown to control the migration of memory T cells to inflamed tissues, as well as stimulate dendritic cell maturation. The chemokine (C-C motif) ligand 19 (CCL19/ECL) has been reported to be a specific ligand of this receptor. ESTs have been isolated from blood, embryo, lymph node, and thymus libraries.
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