MA5-14516

antibody from Invitrogen Antibodies

Targeting: TP53 LFS1, p53

Provider product page for MA5-14516

Western blot

Immunocytochemistry

Immunohistochemistry

Flow cytometry

Other assay

Antibody data

Antibody Data
Antigen structure
References [26]
Comments [0]
Validations
Western blot [2]
Immunocytochemistry [8]
Immunohistochemistry [1]
Other assay [9]

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Product number: MA5-14516 - Provider product page
Provider: Invitrogen Antibodies
Product name: p53 Monoclonal Antibody (SP5)
Antibody type: Monoclonal
Antigen: Recombinant full-length protein
Description: MA5-14516 targets p53 in IHC (P), ICC/IF, and WB applications and shows reactivity with human samples. This antibody does not cross-react with mouse in immunofluorescent applications. The MA5-14516 immunogen is recombinant human full-length wild type p53 protein. This antibody was orginally validated as part of a Thermo Scientific Cellomics High Content Screening Kit. The antibody sold separately may have slightly different performance and may need to be further optimized for the best results.
Reactivity: Human
Host: Rabbit
Isotype: IgG
Antibody clone number: SP5
Vial size: 500 µL
Concentration: 0.004 mg/mL
Storage: -20° C, Avoid Freeze/Thaw Cycles

TP53 protein structure - MA5-14516 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence staining of p21 and p53 in A549 cells treated with media only (non-treated) or with 5 µM camptothecin for 20 hours.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence staining of p21 and p53 in A549 cells treated with media only (non-treated) or with 5 µM camptothecin for 20 hours.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of p53 (green) showing staining in the nucleus of HT29 cells. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with a p53 monoclonal antibody (Product # MA5-14516) in 3% BSA-PBS at a dilution of 1:100 and incubated overnight at 4 ºC in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody in PBS at room temperature in the dark. F-actin (red) was stained with a fluorescent red phalloidin and nuclei (blue) were stained with Hoechst or DAPI. Images were taken at a magnification of 60x.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence staining of p21 and p53 in A549 cells treated with media only (non-treated) or with 5 µM camptothecin for 20 hours.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence staining of p21 and p53 in A549 cells treated with media only (non-treated) or with 5 µM camptothecin for 20 hours.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of p53 (green) showing staining in the nucleus of A431 cells. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with a p53 monoclonal antibody (Product # MA5-14516) in 3% BSA-PBS at a dilution of 1:100 and incubated overnight at 4 ºC in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody in PBS at room temperature in the dark. F-actin (red) was stained with a fluorescent red phalloidin and nuclei (blue) were stained with Hoechst or DAPI. Images were taken at a magnification of 60x.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of TP53 was performed using 70% confluent log phase T-47D 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 p53 Monoclonal Antibody (SP5) (Product # MA5-14516) at 1:100 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 ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). 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 40X. magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of p53 (green) in HeLa cells either left untreated (left panel) or treated with 5uM Camptothecin (right panel) for 20 hours. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA (Product # 37525) for 15 minutes at room temperature. Cells were probed with a p53 monoclonal antibody (Product # MA5-14516) at a dilution of 1:50 for at least 1 hour at room temperature, washed with PBS, and incubated with DyLight 488 goat anti-rabbit IgG secondary antibody (Product # 35552) at a dilution of 1:400 for 30 minutes at room temperature. F-Actin (red) was stained with DyLight 554 Phalloidin (Product # 21834) and nuclei (blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ArrayScan or ToxInsight Instrument at 20X magnification.

Supportive validation

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

Supportive validation

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

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIGURE 5 CircLDB2 regulated non-squamous NSCLC development and cisplatin sensitivity in vitro by miR-346. Vector-transduced or circLDB2-infected A549 and H460 cells were transfected with miR-NC mimic or miR-346 mimic. (a) and (b) CCK-8 assay of cell proliferation. (c) Representative images depicting a colony formation assay. (d) Representative images showing a cell apoptosis assay and flow cytometry for cell apoptosis. (e) Wound-healing assay of cell migration. (f) Cell invasion by transwell assay. (g) and (h) transfected cells were exposed to various concentrations of cisplatin, followed by the evaluation of cell viability by CCK-8 assay. (i) and (j) Western blot showing the levels of p53 and PUMA in transfected cells. * p < 0.05

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIGURE 7 LIMCH1 was a functional target of miR-346. (a) and (b) Western blot analysis of LIMCH1 protein level in cells transfected with sh-NC or sh-LIMCH1. A549 and H460 cells were transfected with anti-NC + sh-NC, anti-miR-346 + sh-NC or anti-miR-346 + sh-LIMCH1. (c) and (d) CCK-8 assay of proliferation of transfected cells. (e) Representative images depicting a colony formation assay. (f) Representative images depicting a cell apoptosis assay and cell apoptosis by flow cytometry. (g) Wound-healing assay of cell migration. (h) Transwell assay of cell invasion. (i) and (j) CCK-8 assay for viability of transfected cells under cisplatin exposure. (k) and (l) Western blot showing the levels of p53 and PUMA in transfected cells. * p < 0.05

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIGURE 8 Overexpression of circLDB2 promoted the antitumor effect of cisplatin in vivo. The xenograft tumors were generated by subcutaneous injection of vector-transduced or circLDB2-transduced A549 cells (2 x 10 6 per mouse, n = 5 per group), and cisplatin treatment was performed twice a week by intraperitoneal injection. Mice were terminated at day 28 after cell implantation. (a) Representative images of the xenograft tumors. (b) Growth curves of the excised tumors. (c) Tumor average weight of the xenograft tumors. (d) Western blot showing the levels of LIMCH1, p53, and PUMA in the excised tumors. * p < 0.05

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: FIGURE 3 CircLDB2 overexpression regulated cell proliferation, apoptosis, migration, invasion, and cisplatin sensitivity in vitro. A549 and H460 cells were transduced with negative control lentivirus (vector) or circLDB2 overexpression lentivirus (circLDB2). (a) and (b) CCK-8 assay of proliferation of transduced cells. (c) Representative images depicting a colony formation assay and colony formation assay of colony formation ability. (d) Representative images depicting a cell apoptosis assay and cell apoptosis by flow cytometry. (e) Representative images showing a cell migration assay and cell migration by wound-healing assay. (f) Representative images showing a cell invasion assay and cell invasion by transwell assay. (g) and (h) CCK-8 assay of viability of transduced cells under cisplatin (various doses) exposure. (i) and (j) representative images depicting a western blot assay and western blot showing the levels of p53 and PUMA in transduced cells. * p < 0.05

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: 4 FIGURE Both mutant p53 and KLF5 increase PLA2G16 expression in pancreatic cancer. A, A heatmap showing TP53 mutation and PLA2G16 expression in pancreatic adenocarcinoma (PAAD) cases in The Cancer Genome Atlas (TCGA). B, A violin plot chart comparing PLA2G16 expression between PAAD cases with or without TP53 mutations. C, Flow chart showing the screening process to identify PLA2G16 correlated TFs in TP53 mutant PAAD cases. D and E, Comparison of KLF5 mRNA expression between normal pancreas in GTEx and PAAD in TCGA (D) and between PAAD cases with or without TP53 mutations (E). F, Representative images of KLF5 protein expression in PAAD tissue. Image credit: Human Protein Atlas, from: https://www.proteinatlas.org/ENSG00000102554-KLF5/pathology/pancreatic+cancer . G and H, Plot charts showing the correlation between PLA2G16 and KLF5 mRNA expression in TP53 mutant (G) and TP53 wild-type (H) PAAD cases. I and J, K-M survival analysis of PFS (I) and DSS (J) in PAAD cases in TCGA Pan-Cancer, by median KLF5 expression separation. K and L, Quantitative real-time PCR (qRT-PCR) analysis of PLA2G16 mRNA expression in PANC-1 and MIA-PaCa-2 cells 48 h after lentiviral-mediated TP53 (K) or KLF5 (L) inhibition. M, Western blot analysis of p53, KLF5 and PLA2G16 protein expression 72 h after lentiviral-mediated TP53 (up) or KLF5 (down) inhibition. N and O, qRT-PCR (M) and Western blot assay (O) of PLA2G16 expression in PANC-1 and MIA-PaCa-2 cells with TP53 and KLF5 inhibition (sh TP53 #2 and

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 5 FIGURE Mutant p53 enhances KLF5-induced PLA2G16 transcriptional activation. A, Predicted binding sites of KLF5 in the promoter region of PLA2G16 . B, The promoter activity of the PLA2G16 gene was measured using a dual-luciferase reporter assay. PANC-1 and MIA-PaCa-2 cells were transfected with pGL3-basic or reporter constructs carrying different lengths of the 5'-flanking region of the PLA2G16 promoter as indicated. C-E, KLF5 depletion reduced the activity of the PLA2G16 promoter. PANC-1 and MIA-PaCa-2 cells with or without lentiviral-mediated KLF5 inhibition were transfected reporter constructs carrying pGL3-(-1303/+239) (C), pGL3-(-400/+239) (D) and pGL3-(-100/+239) (E). 24 h later, luciferase activity was determined. F, Western blot assay of KLF5 expression 48 h after lentiviral-mediated overexpression. G-J, 24 h after infection with lenti-KLF5 or vector, PANC-1 and MIA-PaCa-2 cells were transfected reporter constructs carrying pGL3-(-1303/+239) (G-H) or mutant-pGL3-(-1303/+239) with mutant sequences (C to A) of the four binding sites (I, J). 24 h later, luciferase activity was determined. K, Schematic image showing the location of the designed primer sets for ChIP-quantitative polymerase chain reaction (qPCR) assay, by anti-KLF5 immunoprecipitation. L-O, ChIP-qPCR assays were performed using anti-KLF5 (L, M) or anti-TP53 (N, O) and control IgG antibodies in PANC-1 (L, N) and MIA-PaCa-2 (M, O) cells. Fold enrichment of the indicated PLA2G16 promoter segments was calculat

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig 4 Nuclear intensity of key BER proteins. A) Quantification of mean nuclear fluorescence intensity is shown for PARP1 (**** P < 0.0001, MDA-231 to MDA-157, HCC1806, and MDA-468), PAR (**** P < 0.0001, MDA-468 to MDA-157, MDA-231, and HCC1806), and p53 (**** P < 0.0001, MDA-157 to MDA-231, MDA-157 to MDA-468, MDA-157 to HCC1806, MDA-231 to MDA-468, MDA-231 to HCC1806, and MDA-468 to HCC1806). B) Representative fluorescent images used for quantification in A for PARP1, PAR, and p53 for all cell lines tested. Scale bar = 20 mum.