- PA5-54098 - Invitrogen Antibodies WIPI2 antibody

Thomas HE, Zhang Y, Stefely JA, Veiga SR, Thomas G, Kozma SC, Mercer CA
Cell reports 2018 Aug 28;24(9):2404-2417.e8

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of WIPI2 in human cell line NTERA-2 using a WIPI2 Polyclonal Antibody (Product # PA5-54098).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of WIPI2 in mouse cell line NIH-3T3 and rat cell line NBT-II using a WIPI2 Polyclonal Antibody (Product # PA5-54098).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: KD of WIPI2 was achieved by transfecting SH-SY5Y with WIPI2 specific siRNAs (Silencer® select Product # S25100, S25101). Western blot analysis (Fig. a) was performed using membrane enriched cell extracts from the WIPI2 KD cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with WIPI2 Polyclonal Antibody (Product # PA5-54098, 0.4 µg/mL) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25µg/mL, 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 WIPI2. An uncharacterized band (*) was observed at ~59 kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-WIPI2 Rabbit Polyclonal Antibody (Product # PA5-54098) and a 49 kDa band corresponding to WIPI2 was observed across cell lines tested along with an uncharacterized band (*) at ~60 kDa. Membrane enriched cell extracts (30 µg lysate) of A-431 (Lane 1), THP-1 (Lane 2), Jurkat (Lane 3), SH-SY5Y (Lane 4), Raji (Lane 5) and K-562 (Lane 6) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris 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 (0.4 µg/mL) and detected by chemiluminescence Goat Anti-Rabbit IgG Secondary Antibody, HRP conjugate (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).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent staining of WIPI2 in human cell line A-431 shows positivity in cytoplasm & nucleus but excluded from the nucleoli. Samples were probed using a WIPI2 Polyclonal Antibody (Product # PA5-54098).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical staining of WIPI2 in human testis using a WIPI2 Polyclonal Antibody (Product # PA5-54098) shows cytoplasmic positivity in cells in seminiferous ducts and Leydig cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 4. Phenformin Suppresses the Initiation, Amplitude, and Duration ofAutophagy (A) GST-BHMT assay in cells treated for 6 hr with V, RB, P 1 ,RBP 1 , with or without 1 mM adenosine. (B) Bar graph of AMP/ATP ratios in HEK293 cells treated with V, RB,P 1 , RBP 1 , or starved of glucose. (C and D) Immunofluorescence of WIPI2 in HEK293T cells treated for 2hr. Representative confocal images (C) and quantification of WIPI2 puncta (D).Scale bar is 10 mum. (E-G) GST-BHMT time course assay in cells treated with V, RB, orRBP 1.5 , as shown by western blotting (E) and graphs of BHMTfrag/GFP ratios (F) and LC3-II/ACTIN ratios (G).

PA5-54098