Key features and details
- Mouse monoclonal [DCS2.2] to Cyclin D3/CCND3
- Suitable for: Flow Cyt, WB, IHC-P, IP
- Knockout validated
- Reacts with: Human
- Isotype: IgG1
Product nameAnti-Cyclin D3/CCND3 antibody [DCS2.2]
See all Cyclin D3/CCND3 primary antibodies
DescriptionMouse monoclonal [DCS2.2] to Cyclin D3/CCND3
Tested applicationsSuitable for: Flow Cyt, WB, IHC-P, IPmore details
Species reactivityReacts with: Human
Predicted to work with: Mouse, Rat
Recombinant full length protein corresponding to Human Cyclin D3/CCND3.
MELLCCEGTRHAPRAGPDPRLLGDQRVLQSLLRLEERYVPRASYFQCVQR EIKPHMRKML AYWMLEVCEEQRCEEEVFPLAMNYLDRYLSCVPTRKAQ LQLLGAVCMLLASKLRETTPLT IEKLCIYTDHAVSPRQLRDWEVLVLG KLKWDLAAVIAHDFLAFILHRLSLPRDRQALVKK HAQTFLALCATDYT FAMYPPSMIATGSIGAAVQGLGACSMSGDELTELLAGITGTEVDCL RA CQEQIEAALRESLREASQTSSSPAPKAPRGSSSQGPSQTSTPTDVTAIHL
Database link: P30281
- WB: HeLa, K562, HAP1, HEK-293T and Jurkat cell lysates. Flow Cyt: HeLa cells. IHC-P: Human pancreas tissue.
This product was previously labelled as Cyclin D3
Storage instructionsShipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
Storage bufferPreservative: 0.08% Sodium azide
Concentration information loading...
PurityProtein A/G purified
KO cell lines
KO cell lysates
Our Abpromise guarantee covers the use of ab28283 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use 1µg for 106 cells.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|WB||Use at an assay dependent concentration. Predicted molecular weight: 35 kDa.|
|IHC-P||Use at an assay dependent concentration.|
|IP||Use at an assay dependent concentration.|
FunctionRegulatory component of the cyclin D3-CDK4 (DC) complex that phosphorylates and inhibits members of the retinoblastoma (RB) protein family including RB1 and regulates the cell-cycle during G(1)/S transition. Phosphorylation of RB1 allows dissociation of the transcription factor E2F from the RB/E2F complex and the subsequent transcription of E2F target genes which are responsible for the progression through the G(1) phase. Hypophosphorylates RB1 in early G(1) phase. Cyclin D-CDK4 complexes are major integrators of various mitogenenic and antimitogenic signals. Also substrate for SMAD3, phosphorylating SMAD3 in a cell-cycle-dependent manner and repressing its transcriptional activity. Component of the ternary complex, cyclin D3/CDK4/CDKN1B, required for nuclear translocation and activity of the cyclin D-CDK4 complex.
Sequence similaritiesBelongs to the cyclin family. Cyclin D subfamily.
Contains 1 cyclin N-terminal domain.
Cellular localizationNucleus. Cytoplasm. Membrane. Cyclin D-CDK4 complexes accumulate at the nuclear membrane and are then translocated to the nucleus through interaction with KIP/CIP family members.
- Information by UniProt
- CCND 3 antibody
- Ccnd3 antibody
- CCND3_HUMAN antibody
All lanes : Anti-Cyclin D3/CCND3 antibody [DCS2.2] (ab28283) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : CCND3 knockout HeLa cell lysate
Lane 3 : Jurkat cell lysate
Lane 4 : HEK-293 cell lysate
Lysates/proteins at 20 µg per lane.
All lanes : Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) at 1/10000 dilution
Predicted band size: 35 kDa
Observed band size: 35 kDa
Lanes 1-4: Merged signal (red and green). Green - ab28283 observed at 35 kDa. Red - loading control ab52901.
ab28283 Anti-Cyclin D3/CCND3 antibody [DCS2.2] was shown to specifically react with Cyclin D3 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264931 (knockout cell lysate ab257876) was used. Wild-type and Cyclin D3 knockout samples were subjected to SDS-PAGE. ab28283 and Anti-beta Tubulin [EP1331Y] - Microtubule Marker (ab52901) were incubated at room temperature for 2.5 hours at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
Overlay histogram showing HeLa cells stained with ab28283 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab28283, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was a goat anti-mouse DyLight® 488 (IgG; H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.
ab28283 (4µg/ml) staining Cyclin D3/CCND3 in human pancreas, using an automated system (DAKO Autostainer Plus). Using this protocol there is strong nuclear staining.
Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer citrate pH6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: Cyclin D3/CCND3 (KO) knockout HAP1 whole cell lysate (20 µg)
Lane 3: HEK293 whole cell lysate (20 µg)
Lane 4: Jurkat whole cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab28283 observed at 33 kDa. Red - loading control, ab176560, observed at 50 kDa.
ab28283 was shown to specifically recognize CCND3 in wild-type HAP1 cells along with additional cross reactive bands. No bands was observed when CCND3 knockout samples were uexamined. Wild-type and CCND3 knockout samples were subjected to SDS-PAGE. Ab28283 and ab176560 (Rabbit anti alpha Tubulin loading control) were incubated overnight at 4°C at 1 ug/ml and 1/10,000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/20,000 dilution for 1 hour at room temperature before imaging.
All lanes : Anti-Cyclin D3/CCND3 antibody [DCS2.2] (ab28283) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate
Lane 2 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate
Lane 3 : K562 (Human erythromyeloblastoid leukemia cell line) Whole Cell Lysate
Lysates/proteins at 10 µg per lane.
All lanes : Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 35 kDa
Observed band size: 35 kDa
Additional bands at: 75 kDa. We are unsure as to the identity of these extra bands.
Exposure time: 8 minutes
ab28283 has been referenced in 22 publications.
- Zhang W et al. lncRNA MIAT promotes cell invasion and migration in esophageal cancer. Exp Ther Med 19:3267-3274 (2020). PubMed: 32266022
- Chen CY et al. Embryonic Cul4b is important for epiblast growth and location of primitive streak layer cells. PLoS One 14:e0219221 (2019). PubMed: 31260508
- Ding L et al. Perturbed myoepithelial cell differentiation in BRCA mutation carriers and in ductal carcinoma in situ. Nat Commun 10:4182 (2019). PubMed: 31519911
- Rueda EM et al. The Hippo Pathway Blocks Mammalian Retinal Müller Glial Cell Reprogramming. Cell Rep 27:1637-1649.e6 (2019). PubMed: 31067451
- Zhang Y et al. WD repeat and SOCS box containing protein 2 in the proliferation, cycle progression, and migration of melanoma cells. Biomed Pharmacother 116:108974 (2019). PubMed: 31103822