Invitrogen

Phospholamban Monoclonal Antibody (2D12)

Product Image Gallery

Phospholamban Antibody in Immunofluorescence (IF) — Immunofluorescent analysis of Phospholamban (green) showing staining in the cytoplasm of C2C12 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 Phospholamban monoclonal antibody (Product # MA3-922) in 3% BSA-PBS at a dilution of 1:20 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. Actin was stained using Alexa Fluor 554 (red) and nuclei were stained with Hoechst or DAPI (blue). Images were taken at a magnification of 60x.

Phospholamban Antibody in Immunohistochemistry (Paraffin) (IHC (P)) — Immunofluorescent analysis of Phospholamban (green) showing staining in the cytoplasm of mouse heart tissue (right) compared to a negative control without primary antibody (left). Formalin-fixed tissue was 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. Tissue was probed with a Phospholamban monoclonal antibody (Product # MA3-922) in 3% BSA-PBS at a dilution of 1:100 and incubated overnight at 4°C in a humidified chamber. Tissue was washed with PBST and incubated with a DyLight-conjugated secondary antibody in PBS at room temperature in the dark. Nuclei were stained with Hoechst or DAPI (blue). Images were taken at a magnification of 60x.

Phospholamban Antibody in Immunohistochemistry (IHC) — Immunohistochemistry was performed on normal deparaffinized Human skeletal muscle tissues. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:200 with a mouse monoclonal antibody recognizing Phospholamban (Product # MA3-922) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

Phospholamban Antibody in Western Blot (WB) — Western blot analysis of Phospholamban was performed by loading 25 µg of human heart (lane 1) and C2C12 (lane 2) onto an SDS polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked at 4°C overnight. The membrane was probed with a Phospholamban monoclonal antibody (Product # MA3-922) 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 ~25 kDa.

Phospholamban Antibody — Figure 2. PP1s differentially regulate substrate phosphorylation in neonatal cardiac myocytes Western blot analysis for the phosphorylation of PLB at serine 16 and threonine 17, total PLB, and PP1s from neonatal rat cardiac myocytes infected with adenoviruses expressing either beta-gal (Con) or individual PP1 isoform. p-MLC2 was performed using ProQ-Diamond staining method. Quantification was based on individual western blot. N = 4 for each group. * P < 0.05 vs. Con 5 min.

Product Details

Applications

Tested Dilution

Publications

Immunocytochemistry (ICC)

1:10-1:100

View 4 publications 4 publications

Immunofluorescence (IF)

1:10-1:100

View 1 publication 1 publication

Immunohistochemistry (Paraffin) (IHC (P))

1:200

Immunoprecipitation (IP)

Assay dependent

Western Blot (WB)

1:500-1:5000

View 128 publications 128 publications

Immunohistochemistry (Frozen) (IHC (F))

View 1 publication 1 publication

Immunohistochemistry (IHC)

View 5 publications 5 publications

Neutralization (Neu)

View 1 publication 1 publication

Product Specifications

Species

Dog, Chicken, Guinea pig, Human, Mouse, Sheep, Pig, Rabbit, Rat

Published species

Artificial Control, Dog, Guinea pig, Human, Mouse, Pig, Rabbit, Rat, Rodent

Expression System

Mouse / IgG2a

Class

Monoclonal

Type

Antibody

Clone

2D12

Immunogen

Synthetic peptide corresponding to residues D(2) K V Q Y L T R S A I R R A S T I E M P Q Q A R(25) of canine Phospholamban.

Conjugate

Unconjugated

Form

Liquid

Concentration

1 mg/mL

Purification

Protein A

Storage buffer

PBS with 1mg/mL BSA

Contains

0.05% sodium azide

Storage conditions

-20° C, Avoid Freeze/Thaw Cycles

RRID

AB_2252716

Product Specific Information

MA3-922 detects phospholamban (PLB) in rabbit, human, pig, sheep, chicken, canine, guinea porcine, rat, and mouse.

MA3-922 has been successfully used in Western blot, immunofluorescence, immunohistochemistry and immunoprecipitation procedures. By Western blot, this antibody detects a 25 kDa protein under non-reducing conditions representing the pentameric form of PLB and a 5 kDa protein under reducing conditions representing the monomeric form of PLB in canine cardiac extracts. Immunofluorescence staining of PLB in rat papillary muscle tissue with MA3-922 results in diffuse staining of the network sarcoplasmic reticulum (SR). MA3-922 can also be used to increase the calcium uptake in ventricular SR vesicles analogous to the phosphorylation of PLB following adrenergic stimulation. This antibody is recommended for immunofluorescent staining of tissues.

The MA3-922 immunogen is a synthetic peptide corresponding to residues D(2) K V Q Y L T R S A I R R A S T I E M P Q Q A R(25) of canine PLB. This antibody recognizes an epitope between amino acid residues 9-17 of canine PLB.

Target Information

Phospholamban (PLB) is a 52 amino acid phosphoprotein which regulates the calcium pump of cardiac sarcoplasmic reticulum (SR). PLB is an oligomer of five identical subunits each having a cytoplasmic and transmembrane domain. The cytoplasmic domain (residues 1 to 25) contains the phosphorylation sites and is highly basic and readily cleaved by proteases whereas the transmembrane domain (residues 25 to 52) is mostly hydrophobic, protease resistant and stabilizes the pentamer.

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