MA5-27527

antibody from Invitrogen Antibodies

Targeting: METTL3 M6A, MT-A70, Spo8

Provider product page for MA5-27527

Western blot

Other assay

Antibody data

Antibody Data
Antigen structure
References [1]
Comments [0]
Validations
Other assay [4]

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Product number: MA5-27527 - Provider product page
Provider: Invitrogen Antibodies
Product name: METTL3 Monoclonal Antibody (OTI1B7)
Antibody type: Monoclonal
Antigen: Recombinant protein fragment
Reactivity: Human, Mouse, Rat
Host: Mouse
Isotype: IgG
Antibody clone number: OTI1B7
Vial size: 100 µL
Concentration: 1 mg/mL
Storage: -20° C, Avoid Freeze/Thaw Cycles

METTL3 protein structure - MA5-27527 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig 2 DDX5 directly interacts with METTL3. A: Co-IP of DDX5 and METTL3 in HEK293T cells. B: Co-localization of DDX5 and METTL3 in HEK293T cells cultured in slices and co-transfected with Myc-DDX5 and Flag-METTL3 plasmids. After 24 h, slices were fixed, permeabilized, and incubated with rabbit anti-Myc and mouse anti-Flag antibodies followed by Alexa Fluor 546 labeled goat anti-rabbit IgG (H+L), Alexa Fluor 488 labeled goat anti-mouse IgG (H+L). Nuclei were stained with DAPI; cells were observed by LSCM. Scale bars, 5 mum. C: Schematic of full-length METTL3 and its truncated mutants. D: Co-IP of DDX5 and METTL3 mutants in HEK293T cells. Cells were co-transfected with Myc-DDX5 and Flag-METTL3 mutants. After 24 h, cells were lysed and subjected toIP with Flag antibody, and whole-cell lysates and IP were analyzed by western blotting. E: Co-localization of DDX5 and METTL3 mutants in HEK293T cells. Scale bars, 10 mum. F: Schematic of full-length DDX5 and its truncated mutants. G: Co-IP of METTL3 and DDX5 mutants in HEK293T cells co-transfected with Flag-METTL3 and Myc-DDX5 mutants. After 24 h, cells were lysed and underwent IP with Myc antibody. Whole-cell lysates and IP were analyzed by western blotting. H: Co-localization of METTL3 and DDX5 mutants in HEK293T cells cultured and co-transfected with Flag-METTL3 and Myc-DDX5 mutant plasmids. After 24 h, slices underwent IFA. Scale bars, 10 mum.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig 3 DDX5 regulated the m6A writer complex by recruiting METTL3 after VSV infection. (A, B): VSV promoted the interaction of DDX5 and METTL3 in MEFs. MEFs were infected with VSV or treated with Poly (I:C) for 8h and lysed with NP40 lysis buffer. Lysates were subjected to IP with DDX5 antibody or IgG. Whole-cell and IP lysates were analyzed by western blotting. The DDX5-METTL3 interaction complex was quantified by the western blot band density of METTL3/DDX5 of IP lysates. (C, D): Co-localization of DDX5 and METTL3 in MEFs (C) or macrophages (D) after VSV infection. MEFs/macrophages were cultured in slices and transfected with VSV or treated with Poly (I:C) for 8h. Slices were fixed, permeabilized, and incubated with rabbit anti-DDX5 and mouse anti-METTL3 antibodies followed by Alexa Fluor 546 labeled goat anti-rabbit IgG and Alexa Fluor 488 labeled goat anti-mouse IgG. Nuclei were stained with DAPI. Cells were observed by LSCM. Scale bars, 10 mum. (E, F): The interaction of METTL3 and METTL14 in DDX5-knockdown MEFs (E) or enhanced green fluorescent (EGF) tag fused DDX5-overexpressed MEFs (F). MEFs were transfected with different doses of DDX5 siRNA (siNC) for 48 h (E) or with different doses of EGF-DDX5 expression plasmids (control vector) for 24 h. Cells were infected with VSV (MOI = 10) for 8 h and lysed with NP40 lysis buffer, lysates were subjected to IP with rabbit METTL3 antibody or IgG. Whole-cell and IP lysates were analyzed by western blotting. (G, H): METTL3-METTL1

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig 7 DDX5 negatively regulated DHX58-mediated TBK1 and p65 pathways in innate immunity. (A, B): Immunoblot of DHX58-mediated TBK1 and p65 pathways in DDX5-knockdown MEFs (A) or DDX5-expressing MEFs (B). MEFs were transfected with DDX5 siRNA or METTL3 siRNA (siNC) for 48 h (A), or with DDX5 or METTL3 expression (control vector) for 24 h (B); then, cells were infected with VSV for 0, 4, and 6h, lysed, and collected to detect DDX5, METTL3, p-IKKgamma, IKKgamma,p-p65,p65,p-TBK1, TBK1 and p-IRF7 via western blot. (C, D): Nuclear transfer of IRF7 was observed by IFA and CLSM. MEFs were transfected with DDX5-expression plasmids (control vector) for 24 h (C) or with DDX5 siRNA (siNC) for 48 h (D), infected with VSV for 8 h, and then subjected to IFA with anti-IRF7 monoclonal antibody followed by Alexa Fluor 488 F(ab'')2 fragment of goat anti-mouse IgG (H+L). Nuclei were stained with DAPI. Normal MEFs (WT) were used as the control. Slices were observed by LSCM. Scale bars, 20 mum. (E, F): Nuclear transfer of p65 observed by IFA and CLSM. MEFs were transfected with DDX5-expression plasmids (E) or with DDX5 siRNA (F), infected with VSV for 8h, and subjected to IFA with anti-p65 monoclonal antibody followed by Alexa Fluor 488 F (ab'')2 fragment of goat anti-mouse IgG (H+L). Nuclei were stained with DAPI. Normal MEFs (WT) were used as the control. Slices were observed by LSCM. Scale bars, 20 mum.