- PA5-49581 - Invitrogen Antibodies KMT2D antibody

Submitted references The low-complexity domains of the KMT2D protein regulate histone monomethylation transcription to facilitate pancreatic cancer progression.

Li W, Wu L, Jia H, Lin Z, Zhong R, Li Y, Jiang C, Liu S, Zhou X, Zhang E
Cellular & molecular biology letters 2021 Nov 10;26(1):45

Cul4A-DDB1-mediated monoubiquitination of phosphoglycerate dehydrogenase promotes colorectal cancer metastasis via increased S-adenosylmethionine.

Zhang Y, Yu H, Zhang J, Gao H, Wang S, Li S, Wei P, Liang J, Yu G, Wang X, Li X, Li D, Yang W
The Journal of clinical investigation 2021 Nov 1;131(21)

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Histone-lysine N-methyltransferase 2D was performed using 70% confluent log phase A-431 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with KMT2D Polyclonal Antibody (Product # PA5-49581) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790), (1:2000), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Histone-lysine N-methyltransferase 2D was performed using 70% confluent log phase MCF-7 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with KMT2D Polyclonal Antibody (Product # PA5-49581) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790), (1:2000), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 2 LCD knockout strategy and verification in HEK293T and PANC-1 cells. a Schematic representation of LCD knockout strategy using CRSPR-Cas9 technology. LCD1 (located between exon 8 and 11) was deleted with sgRNA primers: two upstream primers (sgRNA-A1/A2) and two downstream primers (sgRNA-B1/B2). This was then verified by genotyping with KMT2D-1-F/R primers. For LCD2 (located between exon 31 and 36), two upstream primers (sgRNA-C1/C2) and two downstream primers (sgRNA-D1/D2) were used to delete LCD2 and then the rest was tested with KMT2D-2-F/R primers. b , c Verification with genotyping assays. In HEK293T and PANC-1 cells, genotyping PCR was performed in transfected cells. The results are consistent with the predictions compared with the control group (vehicle). d , e RT-PCR analyses to measure LCD knockout. lcd1 mRNA expression only significantly decreased in LCD1-deleted cells, while that of lcd2 only in LCD2-deleted cells. The set gene located at the C-terminal of the KMT2D gene was selected as a control (error bars denote standard deviation). f , g Representative images of immunofluorescences and western blotting in HEK293T and PANC-1 cells. After LCD knockout, the KMT2D protein level did not significantly decrease, and KMT2D LCD1 or KMT2D LCD2 was successfully deleted. KMT2D, 593 KDa; LCD1 deletion-KMT2D, 483 KDa; LCD2 deletion-KMT2D, 447 KDa. All the values are represented as means +- SD. Data are representative of three independent experiments

PA5-49581

Antibody data