PA5-67563

antibody from Invitrogen Antibodies

Targeting: SERPINH1 CBP1, CBP2, colligen, HSP47, SERPINH2

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

PA5-67563

Provider

Invitrogen Antibodies

Product name

SERPINH1 Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Reactivity

Human, Mouse, Rat

Host

Rabbit

Isotype

IgG

Vial size

100 µL

Concentration

0.5 mg/mL

Storage

-20°C

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of SERPINH1 in NIH/3T3 cell (40 µg). Sample was loaded onto a 8% SDS-PAGE gel and probed using a SERPINH1 Polyclonal Antibody (Product # PA5-67563) at a 1:200 dilution dilution, followed by incubation with a Goat anti rabbit IgG secondary antibody at a 1:8000 dilution, Exposure time: 5 seconds.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockdown of Serpin H1 was achieved by transfecting Caco-2 with Serpin H1 specific siRNAs (Silencer® select Product # s2485, s2486). Western blot analysis (Fig. a) was performed using Whole cell extracts from the Serpin H1 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with SERPINH1 Polyclonal Antibody (Product # PA5-67563, 1:500 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to Serpin H1.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-SERPINH1 Polyclonal Antibody (Product # PA5-67563) and a 47 kDa band corresponding to Serpin H1 was observed across tested cell lines along with an uncharacterized band (*) at ~45 kDa. Whole cell extracts (40 µg lysate) of NIH:OVCAR-3 (Lane 1), HeLa (Lane 2), Caco-2 (Lane 3), NIH/3T3 (Lane 4), U-937 (Lane 5), Daudi (Lane 6) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:500 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence analysis of Serpin H1 was performed using 70% confluent log phase Caco-2 cells. The cells were fixed with 4% paraformaldehyde for 5 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with SERPINH1 Polyclonal Antibody (Product # PA5-67563) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790), (1:2000 dilution), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasm (endoplasmic reticulum) localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.