MA5-13128

antibody from Invitrogen Antibodies

Targeting: CCNB1 CCNB

Provider product page for MA5-13128

Western blot

Antibody data

Antibody Data
Antigen structure
References [24]
Comments [0]
Validations
Western blot [2]
Immunocytochemistry [1]
Immunohistochemistry [3]
Flow cytometry [1]
Other assay [5]

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Product number: MA5-13128 - Provider product page
Provider: Invitrogen Antibodies
Product name: Cyclin B1 Monoclonal Antibody (V152)
Antibody type: Monoclonal
Antigen: Other
Description: MA5-13128 targets Cyclin B1 in FACS, IHC (P), and WB applications and shows reactivity with Hamster and Human samples.
Antibody clone number: V152
Concentration: 0.1 mg/mL

CCNB1 protein structure - MA5-13128 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on membrane enriched extracts (30 µg lysate) of MCF7 (Lane 1), HeLa (Lane 2), HeLa treated with Nocodazole (250 nm Nocodazole for 12h) (Lane 3), HT-29 (Lane 4), HT-29 treated with Nocodazole (250 nm Nocodazole for 12h) (Lane 5), HCT-116 (Lane6) and HCT-116 treated with Nocodazole (250 nm Nocodazole for 12h) (Lane 7).The blots were probed with Cyclin B1 Mouse monoclonal Antibody (Product # MA5-13128, 2 µg/mL) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjµgate (Product # A28177, 0.4 µg/mL, 1:2500 dilution). A 55 kDa band corresponding to Cyclin B1 was observed and was enhanced upon treatment in cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of Cyclin B1 was achieved by transfecting HeLa cells with Cyclin B1 specific validated siRNA (Silencer® select Product # s2516). Western blot analysis (Fig a) was performed using whole cell lysates from the Cyclin B1 knock down cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Cyclin B1 Mouse monoclonal Antibody (Product # MA5-13128, 2 µg/mL) and Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.4 µg/mL, 1:2500 dilution). Densitometric analysis of this western blot is shown in histogram(Fig b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to Cyclin B1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Cyclin B1 (V152) showing staining in the nucleus and weak cytoplasm of paraffin-embedded human lung squamous carcinoma (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Cyclin B1 Antibody (V152) Mouse Monoclonal Antibody (Product # MA5-13128) diluted in 3% BSA-PBS at a dilution of 1:100 for 1 hour at 37°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Cyclin B1 (V152) showing staining in the cytoplasm and nucleus of paraffin-embedded human tonsil tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Cyclin B1 Antibody (V152) Mouse Monoclonal Antibody (Product # MA5-13128) diluted in 3% BSA-PBS at a dilution of 1:100 for 1 hour at 37°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Cyclin B1 (V152) showing staining in the cytoplasm and nucleus of paraffin-embedded mouse testis tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Cyclin B1 Antibody (V152) Mouse Monoclonal Antibody (Product # MA5-13128) diluted in 3% BSA-PBS at a dilution of 1:100 for 1 hour at 37°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 FOXM1, CENPF, Cyclin B1 and Pin1 mRNA levels correlate in malignant melanoma and associate with poor disease outcome. ( a ) Heatmaps showing relative z -score corrected mRNA expression profiles of Pin1, FOXM1, CENPF, Cyclin B1 and actin in three independent Oncomine data sets. 43 , 44 , 48 Where applicable, the fold difference between normal skin, benign nevi and melanoma are indicated, all of which are significant ( P 0.45; P 2 years or

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3 Pin1 is required for FOXM1 activity. ( a ) Expression of FOXM1 and CENPF is Pin1-dependent. Immunoblot analysis of the indicated proteins from lysates of wild type and Pin1 - /- MEFs synchronized in G0 by serum deprivation or induced to enter cell cycle progression by 10% FCS for the indicated intervals. N =Nocodazole-induced G2/M arrest. ( b ) Pin1 -/- MEFs show reduced FOXM1 expression under basal conditions. Immunocytochemical detection of Pin1 and FOXM1 in proliferating wild type and Pin1 -/- MEFs. The dashed lines indicate the contours of the nuclei. A merged image is shown to compare co-localization. ( c ) The induction of the FOXM1 target genes CENPF and Cylin-B1 by 4-OHT activatable FOXM1-ER is Pin1-dependent. U2OS cells stably expressing FOXM1-ER were transfected with control siRNA (siCtrl) or siRNA against Pin1 (siPin1). After 24 h they were exposed to 2 or 5muM 4-OHT and 24 h later processed for immunoblot analysis of the indicated proteins. Note that the ER-FOXM1 band is not significantly reduced by Pin1 knockdown, indicating stabilization due to the ER fragment.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 Pin1 is required for FOXM1 activity and melanoma proliferation in an endogenous BRAF V600E background. ( a - c ) Colo829 melanoma cells express elevated levels of Pin1, FOXM1, CENPF and Cyclin B1. ( a ) QPCR-based detection of the indicated mRNAs in primary neonatal Human Epidermal Melanocytes (HEMn) and Colo829 melanoma cells. ( b ) Immunoblot detection of the indicated proteins corresponding to the mRNAs interrogated in ( a ). ( c ) Detection of the indicated proteins interrogated in b by immunofluorescence. ( d ) Endogenous Pin1 and FOXM1 interact in Colo829 cells. Co-immunoprecipitation (IP) of Colo829 lysates (Input) using an antibody against FOXM1 or control IgG. The antibody heavy chains show equal amounts of FOXM1 and control IgG antibodies were used. ( e ) Pin1 is essential for FOXM1 expression and activity. Colo829 cells were transfected with a control siRNA (siCtrl) or siRNA against Pin1 (siPin1) and processed for immunoblot analysis of the indicated proteins. ( f ) Prolonged Pin1 depletion represses Colo829 outgrowth. Colo829 were stably transduced with two independent lentivirally delivered shRNAs targeting Pin1 (shPin1) or a control shRNA (shGFP) and colony formation was addressed 7 days later. ** P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5. Cytoplasmic polyadenylation of Sirena1 . ( A ) Sirena1 RNA abundance in RNA-seq data indicates cytoplasmic polyadenylation during meiotic maturation. The plot depicts the ratios of poly(A)-selected RNA-seq data () relative to total RNA-seq data (), positive numbers and negative log 2 values indicate enrichment and depletion of transcripts in poly(A)-selected RNA-seq data, respectively. In blue are labeled dormant mRNAs with a short poly(A) tail in GV oocytes, which undergo polyadenylation during meiotic maturation. Gray labels depict histone and mitochondrial genes underrepresented in poly(A)-selected RNA-seq data. ( B ) Sequence organization and conservation of the 3'end of Sirena1 . Depicted are putative CPE elements ~100 nucleotides from the poly(A) signal in ten rodent genomes. ( C ) Schematic design of reporters for analysis of cytoplasmic polyadenylation. Renilla luciferase reporter (control) was polyadenylated, other reporters were not polyadenylated. ( D ) Relative luciferase activity of microinjected reporters. Reporters were microinjected into GV oocytes, which were either matured for 20 h (MII) or cultured in the presence of milrinone, which prevents resumption of meiosis (GV). Activity was calculated as luciferase activity of each Renilla reporter divided by a co-injected polyadenylated firefly reporter standard. Error bars = SD. ( E ) Western blot analysis of MOS and CCNB1 proteins in fully-grown GV oocytes and MII eggs. ( F ) Relative abundance of Mos

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2. Menadione induced cell cycle arrest at the G2/M phase. (a) Cell cycle analysis by flow cytometry showing a significant increase in the percentage of cells at G2/M phase after the treatment with menadione in all three GC cell lines. (b) Menadione high significantly increased protein expression of Cyclin B1 and p-CDK1, confirming its potential to arrest cell cycle progression at the G2/M phase. (c) Protein expression of Cyclin B1 and p-CDK1 demonstrated by western blot. The control groups are the respective nontreated cell line.