Invitrogen

PSMB5 Polyclonal Antibody

1 Published Figure

5 References

View all (5) PSMB5 antibodies

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Questions & Answers

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PSMB5 Antibody — Figure 3 UK-101 selectively inhibits LMP2 and its catalytic activity via a stable covalent modification. ( A ) Following a 90 min incubation of cells with UK-101, the concentration dependence of UK-101 binding to LMP2 in PC-3 cells was examined via western blotting. The increase in apparent molecular weight of LMP2 indicates its covalent modification by UK-101. No mobility changes in the X subunit of the constitutive proteasome or the LMP7 subunit of the immunoproteasome were observed. ( B ) The stability of the covalent modification of LMP2 with UK-101 was examined following removal of excess-free UK-101 from the system. PC-3 cells were treated with drug or vehicle for 90 min, then washed and given fresh media. Samples were collected at the indicated times and subjected to western blotting. The binding of UK-101 to LMP2 is maintained for at least 48 h. ( C ) Inhibition of the chymotrypsin-like (CT-L) activity of cellular proteasomes. A proteasome activity assay was performed with PC-3 cells treated with increasing concentrations of proteasome inhibitors (UK-101: 1, 10, 50 mu M; epoxomicin or bortezomib: 5, 50, 500 nM) for 2 h. The CT-L activity of total cellular proteasomes was measured using the CT-L luminogenic substrate Suc-Leu-Leu-Val-Tyr-aminoluciferin. Values were obtained in triplicate, and error bars represent standard deviations. ( D ) Cells were treated with inhibitors, lysed, and probed for ubiquitin. Both UK-101 (25 mu M) and epoxomicin (1 mu M) caused an accumul

PSMB5 Antibody in Immunofluorescence (IF) — Immunofluorescent analysis of Proteasome 20S X (green) showing staining in the cytoplasm and nucleus of BAEC cells (right) compared to a negative control without primary antibody (left). Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with a Proteasome 20S X polyclonal antibody (Product # PA1-977) in 3% BSA-PBS at a dilution of 1:100 and incubated overnight at 4 °C in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody in PBS at room temperature in the dark. F-actin (red) was stained with a fluorescent red phalloidin and nuclei (blue) were stained with Hoechst or DAPI. Images were taken at a magnification of 60x.

PSMB5 Antibody in Western Blot (WB) — Western blot analysis of Proteasome 20S X was performed by loading 25 µg of NCI-H1299 (lane 1), Hela (lane 2) and mouse liver (lane 3) cell lysates onto an SDS polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked at 4°C overnight. The membrane was probed with a Proteasome 20S X polyclonal antibody (Product # PA1-977) at a dilution of 1:1000 overnight at 4°C, washed in TBST, and probed with an HRP-conjugated secondary antibody for 1 hr at room temperature in the dark. Chemiluminescent detection was performed using Pierce ECL Plus Western Blotting Substrate (Product # 32132). Results show a band at ~23 kDa.

Product Details

Tested Applications

Dilution

Immunocytochemistry (ICC)

1:10-1:200

Immunofluorescence (IF)

1:10-1:200

Western Blot (WB)

1:500-1:5000

Published Applications

Western Blot (WB)

See 5 publications below

Product Specifications

Species Reactivity

Bovine, Human, Mouse

Published species

Human, Mouse

Host / Isotype

Rabbit / IgG

Class

Polyclonal

Type

Antibody

Immunogen

Synthetic Peptide: C W(243) I R V S S D N V A D L H D K Y S(259)

Conjugate

Unconjugated

Form

Liquid

Concentration

1 mg/mL

Purification

Antigen affinity chromatography

Storage buffer

PBS with 1mg/mL BSA

Contains

0.05% sodium azide

Storage conditions

-20° C, Avoid Freeze/Thaw Cycles

RRID

AB_2172052

Product Specific Information

PA1-977 detects proteasome 20S X from bovine, mouse and human samples.

PA1-977 has been successfully used in Western blot and ICC/IF procedures. By Western blot, this antibody detects an ~23 kDa protein representing proteasome 20S X.

The PA1-977 immunizing peptide corresponds to amino acid residues 243-259 from mouse proteasome 20S X. This sequence is 94% conserved in human and rat. PA1-977 immunizing peptide (Cat. # PEP-172) is available for use in neutralization and control procedures.

Target Information

The proteasome is a multicatalytic proteinase complex with a highly ordered ring-shaped 20S core structure. The core structure is composed of 4 rings of 28 non-identical subunits; 2 rings are composed of 7 alpha subunits and 2 rings are composed of 7 beta subunits. Proteasomes are distributed throughout eukaryotic cells at a high concentration and cleave peptides in an ATP/ubiquitin-dependent process in a non-lysosomal pathway. An essential function of a modified proteasome, the immunoproteasome, is the processing of class I MHC peptides. This gene encodes a member of the proteasome B-type family, also known as the T1B family, that is a 20S core beta subunit in the proteasome. This catalytic subunit is not present in the immunoproteasome and is replaced by catalytic subunit 3i (proteasome beta 8 subunit). Multiple transcript variants encoding different isoforms have been found for this gene.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

Bioinformatics

Protein Aliases: Macropain epsilon chain; MGC118075; Multicatalytic endopeptidase complex epsilon chain; proteasome (prosome, macropain) subunit, beta type, 5; proteasome beta 5 subunit; proteasome beta type subunit 5; proteasome catalytic subunit 3; Proteasome chain 6; Proteasome epsilon chain; Proteasome subunit beta type-5; Proteasome subunit MB1; Proteasome subunit X; proteasome subunit, beta type, 5; PSX large multifunctional protease X; testicular tissue protein Li 153

Gene Aliases: BOS_10201; LMPX; MB1; PSMB5; X

UniProt ID: (Human) P28074, (Bovine) Q32KL2, (Mouse) O55234

Entrez Gene ID: (Human) 5693, (Bovine) 534640, (Mouse) 19173

Function(s)