XFL2 cDNA was cloned into pDisplay vector (Invitrogen) as described previously (Guselnikov et al., 2008 (link)). In addition, the complete coding regions of X. laevis FcRγ.a (AF499689) and FcRγ.b cDNAs (EF431895) were cloned into pAP-Tag5 vector (GenHunter). As a result, these constructions encoded XFL2 receptor with N-terminal HA epitope and FcRγ subunits with c-myc epitope at their C-end. 293T cells were transiently transfected with the plasmids. Transfections were carried out using Unifectin 56 (IBCH, Moscow, Russia) according o the manufacturer’s protocol. After 72 h transfection, the cells were used for immunocytochemistry and flow cytometry. The surface expression of XFL2 was analyzed with FACSCanto II cytometer (BD Bioscience): live cells were stained with mouse monoclonal 12CA5 anti-HA antibody (Abcam) and goat anti-mouse Ig-FITC (BD Bioscience). Intracellular expression of FcRγ subunits was detected using microscope Axioscop 2 plus (Carl Zeiss): transfected cells were smeared on glass slides, fixed with acetone and stained with anti-c-myc 9E10 monoclonal antibodies (Abcam) and goat anti-mouse IgG-Tex-asRed (Molecular Probes).
Mouse monoclonal 12ca5 anti ha antibody
Manufactured by Abcam
The Mouse monoclonal 12CA5 anti-HA antibody is a laboratory reagent used to detect and identify proteins containing the influenza hemagglutinin (HA) epitope tag. It is a mouse-derived monoclonal antibody that binds specifically to the HA tag, which is commonly used to facilitate the detection and purification of recombinant proteins in research applications.
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Lab products found in correlation
Goat anti mouse igg texas red, by Thermo Fisher Scientific (2 mentions)
Fitc goat anti mouse ig, by BD (2 mentions)
Facscanto 2 cytometer, by BD (2 mentions)
Pdisplay vector, by Thermo Fisher Scientific (2 mentions)
Axioscop 2 plus, by Zeiss (2 mentions)
Anti c myc 9e10 monoclonal antibodies, by Abcam (2 mentions)
2 protocols using mouse monoclonal 12ca5 anti ha antibody
1
Cloning and Expression of XFL2 and FcRγ Subunits
2
Cloning and Expression of XFL2 and FcRγ Subunits
Check if the same lab product or an alternative is used in the 5 most similar protocols
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XFL2 cDNA was cloned into pDisplay vector (Invitrogen) as described previously (Guselnikov et al., 2008 (link)). In addition, the complete coding regions of X. laevis FcRγ.a (AF499689) and FcRγ.b cDNAs (EF431895) were cloned into pAP-Tag5 vector (GenHunter). As a result, these constructions encoded XFL2 receptor with N-terminal HA epitope and FcRγ subunits with c-myc epitope at their C-end. 293T cells were transiently transfected with the plasmids. Transfections were carried out using Unifectin 56 (IBCH, Moscow, Russia) according o the manufacturer’s protocol. After 72 h transfection, the cells were used for immunocytochemistry and flow cytometry. The surface expression of XFL2 was analyzed with FACSCanto II cytometer (BD Bioscience): live cells were stained with mouse monoclonal 12CA5 anti-HA antibody (Abcam) and goat anti-mouse Ig-FITC (BD Bioscience). Intracellular expression of FcRγ subunits was detected using microscope Axioscop 2 plus (Carl Zeiss): transfected cells were smeared on glass slides, fixed with acetone and stained with anti-c-myc 9E10 monoclonal antibodies (Abcam) and goat anti-mouse IgG-Tex-asRed (Molecular Probes).
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