Mouse, Rat, Human, African green monkey Predicted to work with:
Recombinant full length protein corresponding to Human Ubiquitin.
Pancreas and Alzheimer’s brain
This antibody stains the periphery of senile plaques and of neurofibrillary tangles in Alzheimer's disease brain, the Lewy bodies in Parkinson's disease brain, and the Mallory bodies in alcoholic liver disease.
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
1/500 - 1/1000.
1/50 - 1/100.
1/50 - 1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. Boil tissue sections in 10mM citrate buffer, pH 6.0 for 10 min followed by cooling at RT for 20 min.
Use at an assay dependent concentration. Predicted molecular weight: 8.5 kDa. Used at a dilution of 1/2000 B16F10 murine melanoma cell line (see Abreview). Predicted molecular weight: 8.5 kDa.
Use at an assay dependent concentration.
Function: Ubiquitin exists either covalently attached to another protein, or free (unanchored). When covalently bound, it is conjugated to target proteins via an isopeptide bond either as a monomer (monoubiquitin), a polymer linked via different Lys residues of the ubiquitin (polyubiquitin chains) or a linear polymer linked via the initiator Met of the ubiquitin (linear polyubiquitin chains). Polyubiquitin chains, when attached to a target protein, have different functions depending on the Lys residue of the ubiquitin that is linked: Lys-6-linked may be involved in DNA repair; Lys-11-linked is involved in ERAD (endoplasmic reticulum-associated degradation) and in cell-cycle regulation; Lys-29-linked is involved in lysosomal degradation; Lys-33-linked is involved in kinase modification; Lys-48-linked is involved in protein degradation via the proteasome; Lys-63-linked is involved in endocytosis, DNA-damage responses as well as in signaling processes leading to activation of the transcription factor NF-kappa-B. Linear polymer chains formed via attachment by the initiator Met lead to cell signaling. Ubiquitin is usually conjugated to Lys residues of target proteins, however, in rare cases, conjugation to Cys or Ser residues has been observed. When polyubiquitin is free (unanchored-polyubiquitin), it also has distinct roles, such as in activation of protein kinases, and in signaling.
Similarity: Belongs to the ubiquitin family.
Contains 3 ubiquitin-like domains.
Western blot - Anti-Ubiquitin antibody (ab7780)Image courtesy of an anonymous Abreview.
ab7780 used in Western Blot. Whole cell lysate prepared from human lung cancer cell lines was loaded at 100000 cells. The gel was run under reduced, denaturing conditions. 5% milk used as the blocking agent. ab7780 used at a 1/1000 dilution, incubated for 16 hours at 4°C in 5% BSA in TBS-Tween (0.1%). The secondary was an HRP conjugated mouse anti-rabbit, used at a 1/10,000 dilution.Specific bands seen : 16, 24, 32, 50, 60, kDa.
Western blot - Anti-Ubiquitin antibody (ab7780)This image is courtesy of an Abreview submitted by Armen Petrosyan
All lanes : Anti-Ubiquitin antibody (ab7780) at 1/2000 dilution
All lanes : Human Panc-1 whole cell lysate
Lysates/proteins at 700 µg per lane.
Secondary All lanes : HRP Donkey anti-rabbit IgG polyclonal at 1/10000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 8.5 kDa
Exposure time: 25 seconds
Ubiquitin western blot of the complexes pulled down with anti-c-Myc antibodies from the lysates of cells treated with DMSO or MG-132. Treatment with the proteasome inhibitor, MG-132, increases ubiquitinated protein; seen as several bands between 55-72 kDa.
Immunocytochemistry/ Immunofluorescence - Anti-Ubiquitin antibody (ab7780)Image courtesy of Armen Petrosyan by Abreview.
ab7780 staining Ubiquitin in Panc-1 cells by Immunocytochemistry/ Immunofluorescence.Cells were fixed in formaldehyde, blocked with 1% serum for 1 hour at 22°C and then incubated with ab7780 at a 1/2000 dilution for 1 hour at 22°C. The secondary was used at a 1/200 dilution.Green - Golgi residental proteins C2GnT-M.Red -Ubiquitin.Blue - nucleus staining by DAPI.
Immunocytochemistry/ Immunofluorescence - Anti-Ubiquitin antibody (ab7780)Zhao A et al., PLoS One., 2013: 8(2): e56203. Fig 4.; doi: 10.1371/journal.pone.0056203
ab7780 staining ubiquitin in cells transfected with FGAMS-EGFP by ICC/IF (immunocytochemistry/immunofluorescence). Cells were fixed with 3.7% methanol-free formaldehyde at 37°C for 15-20 minutes, blocked with 5% goat serum in PBS-T buffer for 30-60 minutes at room temperature. An Alexa Fluor® 594-conjugated Goat anti-rabbit polyclonal was used as the secondary antibody
ab7780 (1/100) staining Ubiquitin in paraffin-embedded brain sections of a patient with Alzheimer's disease.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Ubiquitin antibody (ab7780)This image is courtesy of an anonymous Abreview
ab7780 staining ubiquitin in Mouse kidney tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 1 hour at 25°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with primary antibody (1/90) for 16 hours at 4°C. A HRP-conjugated Goat anti-rabbit polyclonal (1/1000) was used as the secondary antibody.
ab7780 at a dilution of 1/1000, staining Ubiquitin in neurons (Alexa 488 secondary at 1/2000) on 30µm coronal rat brain (ab29475) tissue sections in free floating IHC (see protocol link for detailed description). Image shows neuronal staining observed with [A] 20x objective and [B] 40x objective.
NB: No labeling observed following omission of primary antibody.
Sections were viewed using an Axioplan 2 Imaging microscope (Imaging Associates) fitted with 10x, 20x and 40x Plan-Neofluorobjectives (Zeiss, Germany) and images were taken using a AxioCam Hrm digital camera (Zeiss, Germany) and AxioVision software (Imaging Associates).