PA1-545

antibody from Invitrogen Antibodies

Targeting: PCYT2 ET

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

PA1-545

Provider

Invitrogen Antibodies

Product name

PCYT2 Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

PA1-545 detects ethanolaminephosphate cytidyltransferase (ECT) protein in human, rat, mouse and non-human primate samples.

Concentration

0.4 mg/mL

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of ECT was performed using whole cell lysates and tissue lysate of Caco-2 (Lane 1), Hep G2 (Lane 2) and Mouse heart (Lane 3). The blot was probed with Anti-ECT Rabbit polyclonal Antibody (Product # PA1-545, 2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A ~ 43 kDa band corresponding to ECT was observed across cell lines and tissue lysate tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody using iBind™ Flex Western Starter Kit (Product # SLF2000S). Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence analysis of PCYT2 was performed using 70% confluent log phase Hep G2 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with PCYT2 Polyclonal Antibody (Product # PA1-545) at 5 µg/mL in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). GRP94 (Panel c: red) was stained with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor 555 (Product # A-31572,2 µg/mL). Panel d represents the merged image showing co-localization of PCYT2 with GRP94 in Endoplasmic Reticulum. Panel e represents the control cells showing no signal. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.