Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Cyclin D1 antibody [SP4] - BSA and Azide free (ab239794)

Overview

  • Product name

    Anti-Cyclin D1 antibody [SP4] - BSA and Azide free
    See all Cyclin D1 primary antibodies
  • Description

    Rabbit monoclonal [SP4] to Cyclin D1 - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: Flow Cyt, ICC, IHC-P, ICC/IF, WB, IHC-Pmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human Cyclin D1 (C terminal). The exact sequence is proprietary.

  • Epitope

    C-terminus
  • Positive control

    • WB: Wild-type HAP1 whole cell lysate; A431 whole cell lysate ICC/IF: Neuro-2a and C6 cells FC: MCF-7, NIH/3T3 and C6 cells
  • General notes

    Ab239794 is the carrier-free version of ab16663. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    ab239794 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

Properties

Applications

Our Abpromise guarantee covers the use of ab239794 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Flow Cyt Use at an assay dependent concentration.
ICC Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Detects a band of approximately 36 kDa (predicted molecular weight: 33 kDa).
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Deparaffinization: Deparaffinize slides using xylene or xylene alternative and graded alcohols.Antigen Retrieval: Boil tissue section in 10mM citrate buffer, pH 6.0 for 10 min followed by cooling at room temperature for 20 min.Primary Antibody Incubation: Incubate for 30 minutes at room temperature.Slide Washing: Slides must be washed in between steps. Rinse slides with PBS/0.05% Tween.

 

Target

  • Function

    Essential for the control of the cell cycle at the G1/S (start) transition.
  • Involvement in disease

    Note=A chromosomal aberration involving CCND1 may be a cause of B-lymphocytic malignancy, particularly mantle-cell lymphoma (MCL). Translocation t(11;14)(q13;q32) with immunoglobulin gene regions. Activation of CCND1 may be oncogenic by directly altering progression through the cell cycle.
    Note=A chromosomal aberration involving CCND1 may be a cause of parathyroid adenomas. Translocation t(11;11)(q13;p15) with the parathyroid hormone (PTH) enhancer.
    Defects in CCND1 are a cause of multiple myeloma (MM) [MIM:254500]. MM is a malignant tumor of plasma cells usually arising in the bone marrow and characterized by diffuse involvement of the skeletal system, hyperglobulinemia, Bence-Jones proteinuria and anemia. Complications of multiple myeloma are bone pain, hypercalcemia, renal failure and spinal cord compression. The aberrant antibodies that are produced lead to impaired humoral immunity and patients have a high prevalence of infection. Amyloidosis may develop in some patients. Multiple myeloma is part of a spectrum of diseases ranging from monoclonal gammopathy of unknown significance (MGUS) to plasma cell leukemia. Note=A chromosomal aberration involving CCND1 is found in multiple myeloma. Translocation t(11;14)(q13;q32) with the IgH locus.
  • Sequence similarities

    Belongs to the cyclin family. Cyclin D subfamily.
  • Post-translational
    modifications

    Phosphorylation at Thr-286 by MAP kinases is required for ubiquitination and degradation following DNA damage. It probably plays an essential role for recognition by the FBXO31 component of SCF (SKP1-cullin-F-box) protein ligase complex.
    Ubiquitinated, primarily as 'Lys-48'-linked polyubiquitination. Ubiquitinated by a SCF (SKP1-CUL1-F-box protein) ubiquitin-protein ligase complex containing FBXO4 and CRYAB (By similarity). Following DNA damage it is ubiquitinated by some SCF (SKP1-cullin-F-box) protein ligase complex containing FBXO31. Ubiquitination leads to its degradation and G1 arrest. Deubiquitinated by USP2; leading to stabilize it.
  • Cellular localization

    Nucleus.
  • Information by UniProt
  • Database links

  • Alternative names

    • AI327039 antibody
    • B cell CLL/lymphoma 1 antibody
    • B cell leukemia 1 antibody
    • B cell lymphoma 1 protein antibody
    • B-cell lymphoma 1 protein antibody
    • BCL 1 antibody
    • BCL-1 antibody
    • BCL-1 oncogene antibody
    • BCL1 antibody
    • BCL1 oncogene antibody
    • ccnd1 antibody
    • CCND1/FSTL3 fusion gene antibody
    • CCND1/FSTL3 fusion gene, included antibody
    • CCND1/IGHG1 fusion gene antibody
    • CCND1/IGHG1 fusion gene, included antibody
    • CCND1/IGLC1 fusion gene antibody
    • CCND1/IGLC1 fusion gene, included antibody
    • CCND1/PTH fusion gene antibody
    • CCND1/PTH fusion gene, included antibody
    • CCND1_HUMAN antibody
    • cD1 antibody
    • Cyl 1 antibody
    • D11S287E antibody
    • G1/S specific cyclin D1 antibody
    • G1/S-specific cyclin-D1 antibody
    • Parathyroid adenomatosis 1 antibody
    • PRAD1 antibody
    • PRAD1 oncogene antibody
    • U21B31 antibody
    see all

Images

  • Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
    Lane 2: CCND1 (Cyclin D1) knockout HAP1 whole cell lysate (20 µg)
    Lane 3: A431 whole cell lysate (20 µg)

    Lanes 1 - 3: Merged signal (red and green). Green - ab16663 observed at 34 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab16663 was shown to specifically recognize CCND1 (Cyclin D1) in wild-type HAP1 cells as signal was lost at the expected MW in CCND1 (Cyclin D1) knockout cells. Wild-type and CCND1 (Cyclin D1) knockout samples were subjected to SDS-PAGE. Ab16663 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1/200 dilution and 1/20,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1/20,000 dilution for 1 hour at room temperature before imaging.

    This data was developed using the same antibody clone in a different buffer formulation containing Tris buffered saline, BSA, and sodium azide (ab16663).

  • Immunocytochemistry/ Immunofluorescence analysis of C6 (rat glial tumor glial cell) cells labeling Cyclin D1 with purified ab16663 at 1/50 (5.42µg/ml). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1/200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • IHC image of ab16663 staining Cyclin D1 in rat esophagus formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab16663, 1:100 dilution, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the secondary only control (shown on the inset).
    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • Flow cytometry analysis of MCF-7 (human breast carcinoma) labeling Cyclin D1 with purified ab16663 at 1/30 dilution (9.03 µg/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as a secondary antibody. Isotypecontrol - Rabbit monoclonal IgG (ab172730) (black). Unlableled control - Unlabelled cells (blue).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • Immunocytochemistry/ Immunofluorescence analysis of Neuro-2a (mouse neuroblastoma neuroblast) cells labeling Cyclin D1� with purified ab16663 at 1/50 (5.42µg/ml). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1/200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • Flow cytometry analysis of C6 (rat glioma) labeling Cyclin D1 with purified ab16663 at 1/30 dilution (9.03 µg/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as a secondary antibody. Isotypecontrol - Rabbit monoclonal IgG (ab172730) (black). Unlableled control - Unlabelled cells (blue).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • Flow cytometry analysis of NIH/3T3 (mouse embryo) labeling Cyclin D1 with purified ab16663 at 1/30 dilution (9.03 µg/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as a secondary antibody. Isotypecontrol - Rabbit monoclonal IgG (ab172730) (black). Unlableled control - Unlabelled cells (blue).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • ab16663 staining Cyclin D1 in wild-type HAP1 cells (top panel) and CCND1 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab16663 at 1/250 dilution and ab195889 at 1/250 dilution (shown in pseudocolour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • ab16663 staining Cyclin D1 in MCF7 cells. The cells were fixed with 4% formaldehyde (10 min), permeabilized in 0.1% Triton X-100 for 5 minutes and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab16663 at a working dilution of 1/250 and ab195889, Mouse monoclonal [DM1A] to alpha Tubulin (Alexa Fluor® 594, shown in red) at 1/250 overnight at +4°C, followed by a further incubation at room temperature for 1h with an anti-rabbit AlexaFluor® 488 (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.
    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • Human mantle cell lymphoma stained with ab16663.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • ab16663 staining Cyclin D1 in Human urinary tract tissue sections by Immunohistochemistry (IHC-P - formaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 10% BSA for 30 minutes at room temperature; antigen retrieval was by heat mediation in citrate buffer. Samples were incubated with primary antibody (1/100 in PBS) for 1 hour. An undiluted HRP-conjugated Goat anti-rabbit IgG polyclonal was used as the secondary antibody.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

  • Immunohistochemical analysis of mouse testis tissue, staining Cyclin D1 with ab16663.

    Antigen retrieval was performed via Tris-EDTA buffer. Sections were blocked with 3% BSA and incubated with primary antibody (1/50) overnight at 4°C. An AlexaFluor®594-conjugated secondary antibody was used to detect staining.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab16663).

References

ab239794 has not yet been referenced specifically in any publications.

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