MA1-010

Masuda Y, Shima G, Aiuchi T, Horie M, Hori K, Nakajo S, Kajimoto S, Shibayama-Imazu T, Nakaya K
The Journal of biological chemistry 2004 Oct 8;279(41):42503-15

Hsp90 chaperone activity requires the full-length protein and interaction among its multiple domains.

Johnson BD, Chadli A, Felts SJ, Bouhouche I, Catelli MG, Toft DO
The Journal of biological chemistry 2000 Oct 20;275(42):32499-507

Felts SJ, Owen BA, Nguyen P, Trepel J, Donner DB, Toft DO
The Journal of biological chemistry 2000 Feb 4;275(5):3305-12

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of TRAP1 was achieved by transfecting HeLa with TRAP1 specific siRNAs (Silencer® select Product # s177). Western blot analysis (Fig. a) was performed using Whole cell extracts from the TRAP1 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with TRAP1 Monoclonal Antibody (TRAP1-6) (Product # MA1-010, 1:2,000 ) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000). 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 TRAP1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-TRAP1 Monoclonal Antibody (TRAP1-6) (Product # MA1-010) and a 74kDa band corresponding to TRAP1 was observed across 4 cell lines. Whole cell extracts (30 µg lysate) of HeLa (Lane 1), Hep G2 (Lane 2), HEK-293 (Lane 3), K-562 (Lane 4) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23002) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:2,000) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of TRAP1 in NCI-H460 Cells. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with a TRAP1 monoclonal antibody (Product # MA1-010) at a dilution of 1:200 overnight at 4 C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody (Product # 35503). TRAP1 staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Images were taken at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of TRAP1 in PC-3-M cells using a TRAP1 monoclonal antibody (Product # MA1-010).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry was performed on normal biopsies of deparaffinized Human liver tissue. 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 and probed with a TRAP1 monoclonal antibody (Product # MA1-010) at a dilution of 1:20 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed 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.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 6 Figure TRAP1 interacts with oADP-ribose in mitochondria. Mitochondria isolated from 10 8 HeLaS3 cells were immunoprecipitated using TRAP1 (75 kDa) antibody. 0.5% input, 5% of final wash, 25% post-elute, and 40% of eluted material of IP either with an unrelated IgG antibody (elute control IgG) or with TRAP1 antibody (elute) were probed for oADPR presence using LP96-10. Whereas the high molecular weight signal is visible in the input fraction, this is no longer detected in the TRAP1-IP elute, but the specific signal at 75 kDa (arrow). * denotes signals from mouse IgG used for IP. IP, immunoprecipitation; PARP, poly(ADP-ribose)polymerase

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunoprecipitation of TRAP1 using Product # MA1-010 visualized by Coomassie Blue staining.