Ubiquitin Antibody (Ubi-1) - BSA Free

Novus Biologicals | Catalog # NB300-130

Novus Biologicals
100% Guarantee

Key Product Details

Validated by

Biological Validation

Species Reactivity

Validated:

Human, Mouse, Rat, Porcine, Bovine, C. elegans, Chicken, Drosophila, Equine, Monkey, Plant, Zebrafish

Cited:

Human, Mouse, Rat, Porcine, Insect - Drosophila, Plant

Applications

Validated:

Immunohistochemistry, Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot, Immunoblotting, Immunocytochemistry/ Immunofluorescence

Cited:

Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot, Immunoblotting, Flow Cytometry, Immunocytochemistry/ Immunofluorescence, IF/IHC

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 Clone # Ubi-1

Format

BSA Free
View all Ubiquitin Primary Antibodies »

Product Specifications

Immunogen

Purified bovine erythrocyte ubiquitin cross-linked with glutaraldehyde to KLH

Reactivity Notes

Plant reactivity reported in scientific literature (PMID: 19147500).

Specificity

Recognizes polyubiquitin chains much more strongly than monoubiquitinated molecules or free ubiquitin. Specifically recognizes ubiquitinated cytoplasmic inclusion bodies.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG1

Theoretical MW

8.5 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for Ubiquitin Antibody (Ubi-1) - BSA Free

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Ubiquitin-Antibody-Ubi-1-Western-Blot-NB300-130-img0014.jpg
Immunocytochemistry/ Immunofluorescence: Ubiquitin Antibody (Ubi-1) [NB300-130]

Immunocytochemistry/ Immunofluorescence: Ubiquitin Antibody (Ubi-1) [NB300-130]

Ubiquitin-Antibody-Ubi-1-Immunocytochemistry-Immunofluorescence-NB300-130-img0013.jpg
Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] - Analysis of HEK293 cell lysates using mouse mAb Ubiquitin, dilution 1:1000 (Green). [1] protein standard (Red), [2] cells maintained in normal medium, [3] cells treated with 10uM of proteasome inhibitor lactacystin (Lc) for 16hrs. Lysed cells were electrophoresed on 4-20% SDS-PAGE and transferred to PVDF membranes. The smear detected above the 200kDa standard represent accumulation of ubiquitinated proteins in proteasome inhibitor-Lc treated cells. The prominent band at 8kDa corresponds to monoubiquitin. Rabbit pAb to HSP60, dilution 1:5000 (Red) was used as a loading control.
Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130]

Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130]

Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130] - FFPE section of cerebral cortex of an Alzheimer patient processed with Ubiquitin antibody using HRP/DAB (Brown) and also stained with haemotoxylin (Blue). A typical flame shaped tangle is seen in a pyramidal neuron in the center and is surrounded by some dystrophic neurites, also strongly ubiquitin positive. Both are commonly seen in cortical and hippocampal Alzheimer brain sections and are typical for this disease, but are rare or absent in healthy brain.
Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130]

Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130]

Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130] - Staining of Ubiquitin in hippocampal tissue from an Alzheimer patient.
Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Ubiquitin-Antibody-Ubi-1-Western-Blot-NB300-130-img0015.jpg
Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130]

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] - Western blot of Ubiquitin expression on (Lane 1) poly-ubiquitin (lys63 linked), (Lane 2) pure ubiquitin and crude homogenates of adult rat, (Lane 3) cortex, (Lane 4) cerebellum and (Lane 5) brain stem.
Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130]

Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130]

Immunohistochemistry-Paraffin: Ubiquitin Antibody (Ubi-1) [NB300-130] - IHC staining of Ubiquitin in human rectal cancer using DAB with hematoxylin counterstain.
Ubiquitin Antibody (Ubi-1)

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] -

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] - In Vivo Applications of Affimers(A) HA-NleL 293 T-Rex cells were induced with 1 μg/mL doxycycline for 12 hr or left untreated. Whole-cell lysate (WCL) blots are shown for actin, HA(-NleL), Ub, & K6 chains. Western blots with the K6 affimer are also shown after Ub enrichment using TUBEs.(B) TUBE-PD of HEK293 cells after 1 hr of MG132 (10 μM) without further treatment (−) or with additional UV (40 J/m2) or CCCP treatment (10 μM for 1 hr) & subsequently blotted with the K6 affimer. Input controls are shown for total Ub & actin & gamma H2AX. The relative signal increase from two experiments is shown below the respective lanes.(C) Expression of WT or catalytically inactive (C431S) Parkin in HeLa Flp-In cells was induced with 0.2 μg/mL doxycycline for 16 hr. Mitochondria were depolarized with O/A for 2 hr. WCL inputs are shown for total Ub, expressed Parkin, TOM20, & actin. The TUBE-PD was also blotted using the K6 affimer & total Ub.(D) Confocal fluorescence microscopy images of cells as in (B) stained with K6 affimer (green), TOM20 (red), & DAPI (blue). Cells & a magnified area are outlined in white. Scale bars correspond to 20 μm.See also Figure S5. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/28943312), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
Ubiquitin Antibody (Ubi-1)

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] -

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] - In Vivo Applications of Affimers(A) HA-NleL 293 T-Rex cells were induced with 1 μg/mL doxycycline for 12 hr or left untreated. Whole-cell lysate (WCL) blots are shown for actin, HA(-NleL), Ub, & K6 chains. Western blots with the K6 affimer are also shown after Ub enrichment using TUBEs.(B) TUBE-PD of HEK293 cells after 1 hr of MG132 (10 μM) without further treatment (−) or with additional UV (40 J/m2) or CCCP treatment (10 μM for 1 hr) & subsequently blotted with the K6 affimer. Input controls are shown for total Ub & actin & gamma H2AX. The relative signal increase from two experiments is shown below the respective lanes.(C) Expression of WT or catalytically inactive (C431S) Parkin in HeLa Flp-In cells was induced with 0.2 μg/mL doxycycline for 16 hr. Mitochondria were depolarized with O/A for 2 hr. WCL inputs are shown for total Ub, expressed Parkin, TOM20, & actin. The TUBE-PD was also blotted using the K6 affimer & total Ub.(D) Confocal fluorescence microscopy images of cells as in (B) stained with K6 affimer (green), TOM20 (red), & DAPI (blue). Cells & a magnified area are outlined in white. Scale bars correspond to 20 μm.See also Figure S5. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/28943312), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
Ubiquitin Antibody (Ubi-1)

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] -

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] - HUWE1 Assembles K6 Chains In Vitro & In Vivo(A) Table summarizing proteins identified with the corresponding number peptide-spectrum matches (PSMs) in three replicates of DUB-treated K6 affimer pull-downs.(B) In vitro assembly reaction of the HECT E3 HUWE1 with Ub WT, Ub K6R, Ub K11R, & Ub K48R on Coomassie. Arrows indicate K6 diUb.(C) Linkage composition of HUWE1-generated diUb after 1 hr as determined by AQUA MS.(D) AQUA-derived total cellular chain composition of HUWE1−/− HeLa & parental cells after TUBE-based enrichment. Error bars indicate mean ± SD from n = 3. ∗p < 0.05, according to a two-tailed Student’s t test. N.S., not significant.(E) TUBE-PD from a doxycycline-inducible HUWE1 shRNA Ls174T cell line blotted with the K6-specific affimer, with input controls for actin, total Ub, & HUWE1.(F) K6 affimer pull-down in doxycycline-inducible HUWE1 shRNA Ls174T cells blotted against Mfn2. Cells were left untreated or treated with 10 μg/mL MG132 for 4 hr and/or 1 μg/mL doxycycline for 72 hr. Pull-downs were incubated with 250 nM USP21 as indicated, with a K48 blot to show completeness of the deubiquitination reaction. Input controls are shown for Mfn2, actin, & HUWE1.See also Figure S7. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/28943312), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
Ubiquitin Antibody (Ubi-1)

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] -

Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130] - In Vivo Applications of Affimers(A) HA-NleL 293 T-Rex cells were induced with 1 μg/mL doxycycline for 12 hr or left untreated. Whole-cell lysate (WCL) blots are shown for actin, HA(-NleL), Ub, & K6 chains. Western blots with the K6 affimer are also shown after Ub enrichment using TUBEs.(B) TUBE-PD of HEK293 cells after 1 hr of MG132 (10 μM) without further treatment (−) or with additional UV (40 J/m2) or CCCP treatment (10 μM for 1 hr) & subsequently blotted with the K6 affimer. Input controls are shown for total Ub & actin & gamma H2AX. The relative signal increase from two experiments is shown below the respective lanes.(C) Expression of WT or catalytically inactive (C431S) Parkin in HeLa Flp-In cells was induced with 0.2 μg/mL doxycycline for 16 hr. Mitochondria were depolarized with O/A for 2 hr. WCL inputs are shown for total Ub, expressed Parkin, TOM20, & actin. The TUBE-PD was also blotted using the K6 affimer & total Ub.(D) Confocal fluorescence microscopy images of cells as in (B) stained with K6 affimer (green), TOM20 (red), & DAPI (blue). Cells & a magnified area are outlined in white. Scale bars correspond to 20 μm.See also Figure S5. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/28943312), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Applications for Ubiquitin Antibody (Ubi-1) - BSA Free

Application
Recommended Usage

Immunocytochemistry/ Immunofluorescence

1:2000

Immunohistochemistry

1:2000

Immunohistochemistry-Frozen

1:2000

Immunohistochemistry-Paraffin

1:2000

Western Blot

1:1000-1:2000
Application Notes
This Ubiquitin Antibody (Ubi-1) is useful for Western blot, ELISA, Immunocytochemistry/Immunofluorescence, and Immunohistochemistry on both paraffin-embedded and frozen sections. In Western blot a band can be seen at 8.5 kDa. This antibody is excellent for the detection of ubiquitinated inclusions seen in human neurodegenerative diseases such as the neurofibrillary tangles of Alzheimer's disease. Use in immunoblotting reported in scientific literature (PMID: 31727601).

Reviewed Applications

Read 4 reviews rated 4.3 using NB300-130 in the following applications:

Formulation, Preparation, and Storage

Purification

Affinity purified

Formulation

50% PBS, 50% glycerol

Format

BSA Free

Preservative

0.035% Sodium Azide

Concentration

1 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: Ubiquitin

Ubiquitin is a highly conserved protein of about 8.5 kDa molecular weight, which has a normal role in the targeting of proteins for proteolytic degradation. To perform this function, the protein to be degraded is first covalently attached to the C-terminus of ubiquitin, and the ubiquitinated complex is then recognized by a complex of degradative enzymes. Interestingly, ubiquitin also becomes covalently bonded to many types of pathological inclusions, which appear to be resistant to normal degradation. Therefore, ubiquitin antibodies are very useful for studies of these inclusions. For example, the neurofibrillary tangles and paired helical filaments diagnostic of Alzheimer's disease, Lewy bodies seen in Parkinson's disease, and Pick bodies found in Pick's disease are all heavily ubiquitinated and can be readily visualized with ubiquitin antibodies.

Alternate Names

UBB

Entrez Gene IDs

7314 (Human); 7316 (Human)

Gene Symbol

UBB

UniProt

P0CG47, P0CG48 (Human)

Additional Ubiquitin Products

Product Documents for Ubiquitin Antibody (Ubi-1) - BSA Free

Certificate of Analysis

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Product Specific Notices for Ubiquitin Antibody (Ubi-1) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Citations for Ubiquitin Antibody (Ubi-1) - BSA Free

Customer Reviews for Ubiquitin Antibody (Ubi-1) - BSA Free (4)

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Customer Images

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  • Ubiquitin Antibody (Ubi-1)
    Name: Anonymous
    Application: Immunohistochemistry-Paraffin
    Sample Tested: Adult heart
    Species: Human
    Verified Customer | Posted 08/25/2021
    Ubiquitin Antibody in heart sample.
    Ubiquitin Antibody (Ubi-1) - BSA Free NB300-130
  • Ubiquitin Antibody (Ubi-1)
    Name: Anonymous
    Application: Western Blot
    Sample Tested: Mouse skeletal muscle homogenate
    Species: Mouse
    Verified Customer | Posted 03/27/2018
    Western Blot: Ubiquitin Antibody (Ubi-1) [NB300-130SS] - Total protein from Mouse skeletal muscle homogenate, separated on a 4-12% gel by SDS-PAGE, transferred to nitrocellulose membrane and blocked in 5% non-fat milk for 1h at room temperature. The membrane was probed with anti-Myogenin 1:1000 in non-fat milk.
    Ubiquitin Antibody (Ubi-1) - BSA Free NB300-130
  • Ubiquitin Antibody (Ubi-1)
    Name: Rune Damgaard
    Application: Western Blot
    Sample Tested: 293 whole cell lysate, Hela whole cell lysate, HCT116 whole cell lysate, MEF whole cell lysate, mouse liver lysate and mouse bone marrow-derived macrophage cells
    Species: Human and Mouse
    Verified Customer | Posted 08/04/2017
    Works well for WB on total cell lysates (boiling the membrane (nitrocellulose) can increase signal drastically if it is low, e.g. when blotting on primary cells/tissues), and it works really well for WB on Ubiquitin pulldowns, e.g. after a TUBE (tandem ubiquitin-binding entity) pulldown.
  • Name: Anonymous
    Application: Immunofluorescence
    Sample Tested: primary mouse neuronal culture
    Species: Mouse
    Verified Customer | Posted 05/27/2009

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Protocols

View specific protocols for Ubiquitin Antibody (Ubi-1) - BSA Free (NB300-130):


Materials

1) 1 Phosphate buffered saline (pH 7.6): NaCl 137mmol/L, KCl 2.7mmol/L, Na2HPO4 4.3mmol/L, KH2PO4 1.4 mmol/L
2) Citrate buffer, 0.01 M, pH6.0, Sodium Citrate 3g, Citric acid 0.4g
3) 3% Hydrogen peroxide
4) Primary antibody
5) Blocking serum (normal serum)
6) Biotinylated secondary antibody
7) DAB staining kit

Methods

1. Dewax and hydration of slides using xylene and EtOH:
Dry slides for 20 min in a 60 C oven
Add Xylene, 2 x 10 min
100%, 95%, 80%, and 70% EtOH, 5 min each EtOH concentration
Rinse in PBS, 5'

2 Antigen retrieval method (only for paraffin slides)
1a. High-pressure antigen retrieval procedure (recommended method)
Place slides in a glass slide holder (ensure that the slide holder is completely filled with slides, slides without sections if necessary, to ensure even heating. The entire slide holder is immersed in 1000 ml of Citrate buffer (0.01M, pH6.0) within a pressure cooker
Once steam is produced, and ONLY when steam is visible, from the pressure cooker (usually 15-20 min), the required high-pressure will have been reached, and slides will be incubated for 2 min.
Turn off heat, and allow buffer and slides to cool to room temperature
Slides are then rinsed in PBS for 5 minutes
2. Add 3% hydrogen peroxide solution, 10'at RT, then PBS, 3X5'
3. Normal blocking serum, 20'at RT
4. Incubate with Primary Ab, 4C overnight or 1.5 hours at 37C
5. Rinse with PBS, 3 X 5' each rinse
6. Add Biotin-conjugated second antibody, 10'at RT
7. Rinse with PBS, 3 X 5' each rinse
8. Add Streptavidin-Peroxidase, 10'at RT
9. Rinse with PBS, 3 X 5' each rinse
10. Staining with DAB solution, 2-5'under microscope
11. Stop the reaction by washing in tap water
12. Counterstain in Haematoxylin for 3-5 minutes
13. 75%, 80%, 95% and 100% ethanol, 5x2', xylene 2 x 10'

Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.

FAQs for Ubiquitin Antibody (Ubi-1) - BSA Free

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  • Q: Regarding the WB image on datasheet of NCX1 antibody (6H2) (NB300-127), can you confirm if lane 2 is a rabbit heart sample? In addition, can you provide the dilution fold and loading mount used to obtain this image? Lastly, would you happen to know if t

    A: A protein stabilizer has not been added to the antibody NB300-130. However, note that this antibody is supplied as ascites fluid, not as a purified antibody. If you are not familiar with this format, the following descriptions will be useful: Ascites fluid is a complex solution containing multiple proteins and background antibodies as well as the antibody of interest. Ascites contains the antibody produced by the hybridoma cells as well as many other proteins, including background antibodies produced by the mice and albumin. Antibody concentrations in ascites are typically high, but an accurate concentration of the target antibody cannot be readily determined. Therefore, unfortunately you cannot determine the concentration of the ubiquitin antibody within NB300-130 as sold. This is why this product's pack sizes are volumes rather than masses, and why we state on our datasheet: 'This product is unpurified. The exact concentration of antibody is not quantifiable.' All I can suggest is that you carry out antigen affinity chromatography of the antibody, and then determine its concentration.

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