Phospho-Retinoblastoma (Ser780) Antibody - #AF3103

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Product: Phospho-Retinoblastoma (Ser780) Antibody
Catalog: AF3103
Description: Rabbit polyclonal antibody to Phospho-Retinoblastoma (Ser780)
Application: WB IHC IF/ICC
Reactivity: Human, Mouse, Rat
Prediction: Pig, Bovine, Horse, Sheep, Rabbit, Dog, Chicken, Xenopus
Mol.Wt.: 106kDa; 106kD(Calculated).
Uniprot: P06400
RRID: AB_2834540

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Related Downloads
MSDS
Data Sheet
COA
Protocols
WB Handbook
IHC Protocol
IF/ICC Protocol

Product Info

Source:
Rabbit
Application:
WB 1:500-1:2000, IHC 1:50-1:200, IF/ICC 1:100-1:500
*The optimal dilutions should be determined by the end user.
*Tips:

WB: For western blot detection of denatured protein samples. IHC: For immunohistochemical detection of paraffin sections (IHC-p) or frozen sections (IHC-f) of tissue samples. IF/ICC: For immunofluorescence detection of cell samples. ELISA(peptide): For ELISA detection of antigenic peptide.

Reactivity:
Human,Mouse,Rat
Prediction:
Pig(100%), Bovine(100%), Horse(91%), Sheep(100%), Rabbit(100%), Dog(100%), Chicken(91%), Xenopus(90%)
Clonality:
Polyclonal
Specificity:
Phospho-Retinoblastoma (Ser780) Antibody detects endogenous levels of Retinoblastoma only when phosphorylated at Serine 780.
RRID:
AB_2834540
Cite Format: Affinity Biosciences Cat# AF3103, RRID:AB_2834540.
Conjugate:
Unconjugated.
Purification:
The antibody is from purified rabbit serum by affinity purification via sequential chromatography on phospho-peptide and non-phospho-peptide affinity columns.
Storage:
Rabbit IgG in phosphate buffered saline , pH 7.4, 150mM NaCl, 0.02% sodium azide and 50% glycerol. Store at -20 °C. Stable for 12 months from date of receipt.
Alias:

Fold/Unfold

Exon 17 tumor GOS561 substitution mutation causes premature stop; GOS563 exon 17 substitution mutation causes premature stop; OSRC; Osteosarcoma; p105-Rb; P105RB; PP105; pp110; PPP1R130; pRb; Prepro retinoblastoma associated protein; Protein phosphatase 1 regulatory subunit 130; Rb; RB transcriptional corepressor 1; RB_HUMAN; RB1; RB1 gene; Retinoblastoma 1; Retinoblastoma suspectibility protein; Retinoblastoma-associated protein;

Immunogens

Immunogen:
Uniprot:

P06400(RB_HUMAN) >>Visit HPA database.

Gene(ID):
RB1(5925)
Expression:
P06400 RB_HUMAN:

Expressed in the retina. Expressed in foreskin keratinocytes (at protein level) (PubMed:20940255).

Description:
Retinoblastoma (RB) is an embryonic malignant neoplasm of retinal origin. It almost always presents in early childhood and is often bilateral. Spontaneous regression ('cure') occurs in some cases.
Sequence:
MPPKTPRKTAATAAAAAAEPPAPPPPPPPEEDPEQDSGPEDLPLVRLEFEETEEPDFTALCQKLKIPDHVRERAWLTWEKVSSVDGVLGGYIQKKKELWGICIFIAAVDLDEMSFTFTELQKNIEISVHKFFNLLKEIDTSTKVDNAMSRLLKKYDVLFALFSKLERTCELIYLTQPSSSISTEINSALVLKVSWITFLLAKGEVLQMEDDLVISFQLMLCVLDYFIKLSPPMLLKEPYKTAVIPINGSPRTPRRGQNRSARIAKQLENDTRIIEVLCKEHECNIDEVKNVYFKNFIPFMNSLGLVTSNGLPEVENLSKRYEEIYLKNKDLDARLFLDHDKTLQTDSIDSFETQRTPRKSNLDEEVNVIPPHTPVRTVMNTIQQLMMILNSASDQPSENLISYFNNCTVNPKESILKRVKDIGYIFKEKFAKAVGQGCVEIGSQRYKLGVRLYYRVMESMLKSEEERLSIQNFSKLLNDNIFHMSLLACALEVVMATYSRSTSQNLDSGTDLSFPWILNVLNLKAFDFYKVIESFIKAEGNLTREMIKHLERCEHRIMESLAWLSDSPLFDLIKQSKDREGPTDHLESACPLNLPLQNNHTAADMYLSPVRSPKKKGSTTRVNSTANAETQATSAFQTQKPLKSTSLSLFYKKVYRLAYLRLNTLCERLLSEHPELEHIIWTLFQHTLQNEYELMRDRHLDQIMMCSMYGICKVKNIDLKFKIIVTAYKDLPHAVQETFKRVLIKEEEYDSIIVFYNSVFMQRLKTNILQYASTRPPTLSPIPHIPRSPYKFPSSPLRIPGGNIYISPLKSPYKISEGLPTPTKMTPRSRILVSIGESFGTSEKFQKINQMVCNSDRVLKRSAEGSNPPKPLKKLRFDIEGSDEADGSKHLPGESKFQQKLAEMTSTRTRMQKQKMNDSMDTSNKEEK

Predictions

Predictions:

Score>80(red) has high confidence and is suggested to be used for WB detection. *The prediction model is mainly based on the alignment of immunogen sequences, the results are for reference only, not as the basis of quality assurance.

Species
Results
Score
Pig
100
Bovine
100
Sheep
100
Dog
100
Rabbit
100
Horse
91
Chicken
91
Xenopus
90
Zebrafish
0
Model Confidence:
High(score>80) Medium(80>score>50) Low(score<50) No confidence

PTMs - P06400 As Substrate

Site PTM Type Enzyme
Phosphorylation
T5 Phosphorylation P11802 (CDK4)
S37 Phosphorylation
K63 Ubiquitination
K65 Ubiquitination
K94 Ubiquitination
K136 Ubiquitination
T140 Phosphorylation
K143 Ubiquitination
S149 Phosphorylation
S163 Phosphorylation
S230 Phosphorylation P24941 (CDK2)
Y239 Phosphorylation
T241 Phosphorylation
S249 Phosphorylation P24941 (CDK2) , P11802 (CDK4) , P05771 (PRKCB) , P06493 (CDK1)
T252 Phosphorylation P06493 (CDK1) , P05771 (PRKCB) , P24941 (CDK2) , P11802 (CDK4)
K265 Ubiquitination
K279 Ubiquitination
K289 Ubiquitination
K319 Ubiquitination
Y321 Phosphorylation
Y325 Phosphorylation
K327 Ubiquitination
K341 Ubiquitination
T345 Phosphorylation
S347 Phosphorylation
S350 Phosphorylation
T353 Phosphorylation
T356 Phosphorylation P24941 (CDK2) , P11802 (CDK4)
K359 Ubiquitination
S360 Phosphorylation
T373 Phosphorylation P06493 (CDK1) , P24941 (CDK2) , P11802 (CDK4)
K420 Ubiquitination
K427 Acetylation
K427 Ubiquitination
K432 Ubiquitination
Y454 Phosphorylation
K537 Ubiquitination
K548 Acetylation
S567 Phosphorylation Q16539 (MAPK14) , P24941 (CDK2)
K574 Ubiquitination
T583 Phosphorylation
S588 Phosphorylation
T601 Phosphorylation
Y606 Phosphorylation
S608 Phosphorylation P24941 (CDK2) , P11802 (CDK4)
S612 Phosphorylation O96017 (CHEK2) , O14757 (CHEK1) , Q00534 (CDK6) , P11802 (CDK4) , P24941 (CDK2)
S618 Phosphorylation
S624 Phosphorylation
T625 Phosphorylation
K640 Ubiquitination
K720 Sumoylation
Y771 Phosphorylation
S773 Phosphorylation
T774 Phosphorylation
R775 Methylation
T778 Phosphorylation
S780 Phosphorylation P11802 (CDK4) , P30279 (CCND2) , P17612 (PRKACA) , Q00534 (CDK6) , Q96GD4 (AURKB) , P05771 (PRKCB)
R787 Methylation
S788 Phosphorylation Q00534 (CDK6) , P11802 (CDK4)
Y790 Phosphorylation
K791 Ubiquitination
S794 Phosphorylation
S795 Phosphorylation P05771 (PRKCB) , P11802 (CDK4) , P50750 (CDK9) , Q00534 (CDK6) , Q00535 (CDK5)
R798 Methylation
Y805 Phosphorylation P00519 (ABL1)
S807 Phosphorylation P24941 (CDK2) , P50750 (CDK9) , Q00534 (CDK6) , P11802 (CDK4) , Q00526 (CDK3) , P06493 (CDK1) , Q00535 (CDK5)
K810 Methylation
K810 Ubiquitination
S811 Phosphorylation Q00534 (CDK6) , P06493 (CDK1) , P24941 (CDK2) , P11802 (CDK4) , Q00526 (CDK3) , P50750 (CDK9) , Q00535 (CDK5) , Q13131 (PRKAA1)
Y813 Phosphorylation
K814 Ubiquitination
S816 Phosphorylation
T821 Phosphorylation P24941 (CDK2) , Q00534 (CDK6) , P05771 (PRKCB)
T823 Phosphorylation
K824 Ubiquitination
T826 Phosphorylation Q00534 (CDK6) , P11802 (CDK4) , P24941 (CDK2)
S829 Phosphorylation
S834 Phosphorylation
S838 Phosphorylation
T841 Phosphorylation
S842 Phosphorylation
K844 Ubiquitination
K847 Ubiquitination
S855 Phosphorylation
R857 Methylation
K860 Methylation
K860 Ubiquitination
S862 Phosphorylation
S866 Phosphorylation
K870 Ubiquitination
K873 Acetylation
K874 Acetylation
S882 Phosphorylation
S895 Phosphorylation
K896 Acetylation
K896 Ubiquitination
K900 Ubiquitination
T905 Phosphorylation
S919 Phosphorylation
K925 Acetylation

Research Backgrounds

Function:

Key regulator of entry into cell division that acts as a tumor suppressor. Promotes G0-G1 transition when phosphorylated by CDK3/cyclin-C. Acts as a transcription repressor of E2F1 target genes. The underphosphorylated, active form of RB1 interacts with E2F1 and represses its transcription activity, leading to cell cycle arrest. Directly involved in heterochromatin formation by maintaining overall chromatin structure and, in particular, that of constitutive heterochromatin by stabilizing histone methylation. Recruits and targets histone methyltransferases SUV39H1, KMT5B and KMT5C, leading to epigenetic transcriptional repression. Controls histone H4 'Lys-20' trimethylation. Inhibits the intrinsic kinase activity of TAF1. Mediates transcriptional repression by SMARCA4/BRG1 by recruiting a histone deacetylase (HDAC) complex to the c-FOS promoter. In resting neurons, transcription of the c-FOS promoter is inhibited by BRG1-dependent recruitment of a phospho-RB1-HDAC1 repressor complex. Upon calcium influx, RB1 is dephosphorylated by calcineurin, which leads to release of the repressor complex (By similarity).

(Microbial infection) In case of viral infections, interactions with SV40 large T antigen, HPV E7 protein or adenovirus E1A protein induce the disassembly of RB1-E2F1 complex thereby disrupting RB1's activity.

PTMs:

Phosphorylated by CDK6 and CDK4, and subsequently by CDK2 at Ser-567 in G1, thereby releasing E2F1 which is then able to activate cell growth. Dephosphorylated at the late M phase. SV40 large T antigen, HPV E7 and adenovirus E1A bind to the underphosphorylated, active form of pRb. Phosphorylation at Thr-821 and Thr-826 promotes interaction between the C-terminal domain C and the Pocket domain, and thereby inhibits interactions with heterodimeric E2F/DP transcription factor complexes. Dephosphorylated at Ser-795 by calcineruin upon calcium stimulation. CDK3/cyclin-C-mediated phosphorylation at Ser-807 and Ser-811 is required for G0-G1 transition. Phosphorylated by CDK1 and CDK2 upon TGFB1-mediated apoptosis (By similarity).

N-terminus is methylated by METTL11A/NTM1 (By similarity). Monomethylation at Lys-810 by SMYD2 enhances phosphorylation at Ser-807 and Ser-811, and promotes cell cycle progression. Monomethylation at Lys-860 by SMYD2 promotes interaction with L3MBTL1.

Acetylated during keratinocyte differentiation. Acetylation at Lys-873 and Lys-874 regulates subcellular localization. Can be deacetylated by SIRT1.

Subcellular Location:

Nucleus.
Note: During keratinocyte differentiation, acetylation by KAT2B/PCAF is required for nuclear localization.

Extracellular region or secreted Cytosol Plasma membrane Cytoskeleton Lysosome Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionSubcellular location
Tissue Specificity:

Expressed in the retina. Expressed in foreskin keratinocytes (at protein level).

Subunit Structure:

Interacts with ATAD5. Interacts with PRMT2, CDK1 and CDK2 (By similarity). The hypophosphorylated form interacts with and sequesters the E2F1 transcription factor. Interacts with heterodimeric E2F/DP transcription factor complexes containing TFDP1 and either E2F1, E2F3, E2F4 or E2F5, or TFDP2 and E2F4. The unphosphorylated form interacts with EID1, ARID3B, KDM5A, SUV39H1, MJD2A/JHDM3A and THOC1. Interacts with the N-terminal domain of TAF1. Interacts with SNW1, AATF, DNMT1, LIN9, LMNA, KMT5B, KMT5C, PELP1, UHRF2 and TMPO-alpha. May interact with NDC80. Interacts with GRIP1 and UBR4. Interacts with ARID4A and KDM5B. Interacts with E4F1 and LIMD1. Interacts with SMARCA4/BRG1 AND HDAC1 (By similarity). Interacts with PSMA3 and USP4. Interacts (when methylated at Lys-860) with L3MBTL1. Interacts with CHEK2; phosphorylates RB1. Interacts with CEBPA. P-TEFB complex interacts with RB1; promotes phosphorylation of RB1. Interacts with RBBP9; the interaction disrupts RB1 binding to E2F1 (By similarity). Interacts with KAT2B/PCAF and EP300/P300 (By similarity).

(Microbial infection) Interacts with adenovirus E1A protein.

(Microbial infection) Interacts with HPV E7 protein.

(Microbial infection) Interacts with SV40 large T antigen.

(Microbial infection) Interacts with human cytomegalovirus/HHV-5 protein UL123.

(Microbial infection) Interacts with molluscum contagiosum virus protein MC007.

Family&Domains:

The Pocket domain binds to the threonine-phosphorylated domain C, thereby preventing interaction with heterodimeric E2F/DP transcription factor complexes.

Belongs to the retinoblastoma protein (RB) family.

Research Fields

· Cellular Processes > Cell growth and death > Cell cycle.   (View pathway)

· Cellular Processes > Cell growth and death > Cellular senescence.   (View pathway)

· Human Diseases > Drug resistance: Antineoplastic > Endocrine resistance.

· Human Diseases > Infectious diseases: Viral > Hepatitis B.

· Human Diseases > Infectious diseases: Viral > Human papillomavirus infection.

· Human Diseases > Infectious diseases: Viral > HTLV-I infection.

· Human Diseases > Infectious diseases: Viral > Epstein-Barr virus infection.

· Human Diseases > Cancers: Overview > Pathways in cancer.   (View pathway)

· Human Diseases > Cancers: Overview > Viral carcinogenesis.

· Human Diseases > Cancers: Specific types > Pancreatic cancer.   (View pathway)

· Human Diseases > Cancers: Specific types > Glioma.   (View pathway)

· Human Diseases > Cancers: Specific types > Prostate cancer.   (View pathway)

· Human Diseases > Cancers: Specific types > Melanoma.   (View pathway)

· Human Diseases > Cancers: Specific types > Bladder cancer.   (View pathway)

· Human Diseases > Cancers: Specific types > Chronic myeloid leukemia.   (View pathway)

· Human Diseases > Cancers: Specific types > Small cell lung cancer.   (View pathway)

· Human Diseases > Cancers: Specific types > Non-small cell lung cancer.   (View pathway)

· Human Diseases > Cancers: Specific types > Breast cancer.   (View pathway)

· Human Diseases > Cancers: Specific types > Hepatocellular carcinoma.   (View pathway)

· Human Diseases > Cancers: Specific types > Gastric cancer.   (View pathway)

References

1). Blockage of EGFR/AKT and mevalonate pathways synergize the antitumor effect of temozolomide by reprogramming energy metabolism in glioblastoma. Cancer communications (London, England), 2023 (PubMed: 37920878) [IF=16.2]

Application: WB    Species: Human    Sample: TBD0220, U-87 MG, and U-87 MG-EGFR-vIII cells

FIGURE 3 EGFR/AKT pathway regulates mitochondrial respiration and proliferation in GBM cells. (A-B) TBD0220 (A), and U-87 MG-EGFR-vIII cells (B) were treated with DMSO or 5 μmol/L MK-2206 for 24 h. The mitochondrial functions were monitored by Seahorse XF Cell Mito Stress test. The OCR, basal respiration, proton leak, and ATP production rates were measured as illustrated. ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001 (independent-sample Student's t-test for TBD0220; one-way ANOVA for U-87 MG). (C) ATP levels in TBD0220, U-87 MG, and U-87 MG-EGFR-vIII cells were analyzed after 24 h of treatments with DMSO or 5 μmol/L of MK-2206. ∗∗P < 0.01, ∗∗∗P < 0.001 (independent-sample Student's t-test for TBD0220; one-way ANOVA for U-87 MG). (D) The cell growth assay for TBD0220, U-87 MG, and U-87 MG-EGFR-vIII lines treated with DMSO, 5 μmol/L MK-2206, or 5 μmol/L MK-2206 plus 50 μmol/L ATP were performed. ∗∗∗P < 0.001 (two-way ANOVA). (E) The colony formation assay of TBD0220, U-87 MG, and U-87 MG-EGFR-vIII lines treated with DMSO or 1 μmol/L MK-2206. (F) Cell cycle distributions were analyzed by flow cytometry in TBD0220, U-87 MG, and U-87 MG-EGFR-vIII cells treated with DMSO or 5 μmol/L MK-2206. (G) Western blotting to show changes in expressions of CDK2, CDK4, CDK6, Cyclin D, RB, p-RB, and GAPDH in TBD0220, U-87 MG, and U-87 MG-EGFR-vIII cells treated with DMSO or MK-2206. Abbreviations: EGFR, epidermal growth factor receptor; AKT, AKT serine/threonine kinase 1; GBM, glioblastoma; ANOVA, analysis of variance; DMSO, dimethyl sulfoxide; CDK2, cyclin-dependent kinase 2; CDK4, cyclin-dependent kinase 4; CDK6, cyclin-dependent kinase 6; OCR, oxygen consumption rate.
2). TCA-phospholipid-glycolysis targeted triple therapy effectively suppresses ATP production and tumor growth in glioblastoma. Theranostics, 2023 (PubMed: 36276638) [IF=12.4]

Application: WB    Species: Human    Sample: TBD0220 and U87MG cells

Figure 3 Reduction in ATP production hinders cell proliferation and contributes to G1/S arrest in GBM. (A-B) The relative viability of TBD0220 (A) and U87MG (B) cells was measured using a CCK-8 kit (n = 3). (C-D) Colony formation assay to detect GBM cell growth (D), and the quantification of colony numbers (C) (n = 3). (E) Cell cycle analysis using flow cytometry after incubation with different treatments like EPIC (20 µM), AA (25 µM), or EPIC (20 µM) + AA (25 µM) for 48 h, and the results are plotted as a histogram (n = 3) (F) Representative western blotting showing the expression of p21 and Rb and their downstream targets. The results were normalized to Tubulin with the control group as 1. Protein expression was quantified by ImageJ. (G) Representative confocal images of CDK6 after the treatment with AA (25 µM), EPIC (20 µM), or EPIC (20 µM) + AA (25 µM) for 48 h (n = 6). (H) The quantitative analysis of fluorescence images of CDK6 (n = 6). All data are shown as the mean values ± SD, and p values are based on one-way or two-way ANOVA. ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05. Scale bar = 20 μm.
3). Combination LSD1 and HOTAIR-EZH2 inhibition disrupts cell cycle processes and induces apoptosis in glioblastoma cells. PHARMACOLOGICAL RESEARCH, 2021 (PubMed: 34246782) [IF=9.3]
4). Metformin as a senostatic drug enhances the anticancer efficacy of CDK4/6 inhibitor in head and neck squamous cell carcinoma. Cell Death & Disease, 2020 (PubMed: 33116117) [IF=9.0]

Application: WB    Species: Human    Sample:

Fig. 2 CDK4/6 inhibitor combined with metformin promoted cell cycle arrest. A, B Flow cytometry results showed that the combination of the CDK4/6 inhibitor LY2835219 (HSC3: 0.1 μM; HSC6: 0.3 μM; Cal27: 1.25 μM) with metformin (HSC3: 10 mM; HSC6: 1 mM; Cal27: 10 mM) induced cell cycle arrest in the G0/G1 phase. C Western blot results showed that LY2835219 upregulated the expression of p16 and p21 and downregulated the expression of pRb. D Western blot results showed that metformin upregulated the expression of p16 and p21 and downregulated the expression of pRb. E Compared with LY2835219 monotherapy, the combination treatment significantly upregulated p21 expression and suppressed p-Rb expression. F Immunohistochemistry results (400×) showed that LY2835219 and metformin synergistically upregulated p21 expression and downregulated pRb expression in HSC6 xenograft tumors. Bar: 100 μm. *P 

Application: IHC    Species: Human    Sample:

Fig. 2 CDK4/6 inhibitor combined with metformin promoted cell cycle arrest. A, B Flow cytometry results showed that the combination of the CDK4/6 inhibitor LY2835219 (HSC3: 0.1 μM; HSC6: 0.3 μM; Cal27: 1.25 μM) with metformin (HSC3: 10 mM; HSC6: 1 mM; Cal27: 10 mM) induced cell cycle arrest in the G0/G1 phase. C Western blot results showed that LY2835219 upregulated the expression of p16 and p21 and downregulated the expression of pRb. D Western blot results showed that metformin upregulated the expression of p16 and p21 and downregulated the expression of pRb. E Compared with LY2835219 monotherapy, the combination treatment significantly upregulated p21 expression and suppressed p-Rb expression. F Immunohistochemistry results (400×) showed that LY2835219 and metformin synergistically upregulated p21 expression and downregulated pRb expression in HSC6 xenograft tumors. Bar: 100 μm. *P 
5). Combination of N, N′-dicyclohexyl-N-arachidonic acylurea and tacrolimus prolongs cardiac allograft survival in mice. Immunology and Cell Biology, 2020 (PubMed: 32162358) [IF=4.0]

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