- PA3-023 - Invitrogen Antibodies PPID antibody

Taherian A, Krone PH, Ovsenek N
Biochemistry and cell biology = Biochimie et biologie cellulaire 2008 Feb;86(1):37-45

Song J, Lu YC, Yokoyama K, Rossi J, Chiu R
The Journal of biological chemistry 2004 Jun 4;279(23):24414-9

Bharadwaj S, Ali A, Ovsenek N
Molecular and cellular biology 1999 Dec;19(12):8033-41

Bharadwaj S, Ali A, Ovsenek N
Molecular and cellular biology 1999 Dec;19(12):8033-41

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of Cyclophilin 40 was achieved by transfecting LNCaP with Cyclophilin 40 specific siRNAs (Silencer® select Product # S10913, S10915). Western blot analysis (Fig. a) was performed using Whole cell extracts from the Cyclophilin 40 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with Cyclophilin 40 Polyclonal Antibody (Product # PA3-023, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20,000 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 Cyclophilin 40.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-Cyclophilin 40 Polyclonal Antibody (Product # PA3-023) and a ~40 kDa band corresponding to Cyclophilin 40 was observed across cell lines and tissues tested. Whole cell lysate (40 µg lysate) of LNCaP (Lane 1), KARPAS 299 (Lane 2), NIH:OVCAR-3 (Lane 3), K-562 (Lane 4), U-2 OS (Lane 5), A-431 (Lane 6), MCF7 (Lane 7), Mouse Brain (Lane 8) and Rat Brain (Lane 9) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX), 10 well. 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 (1:1000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20,000 dilution) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Cyclophilin D was done on 70% confluent log phase A431 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Cyclophilin D Rabbit Polyclonal Antibody (Product # PA3-023) at 1:250 dilution in 0.1% BSA and incubated for 3 hours at room temperature 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 SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d is a merged image showing cytoplasmic localization. Panel e is a no primary antibody control. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Cyclophilin 40 was performed using 70% confluent log phase A549 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.01% Triton™ X-100 for 10 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with Cyclophilin 40 Polyclonal Antibody (Product # PA3-023) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000 dilution), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with Hoechst 33342 (Product # H1399). F-actin (Panel c: Red) was stained with Alexa Fluor™ Plus 647 Phalloidin (Product # A30107, 1:2000 dilution). Panel d represents the merged image showing Nucleus and cytoplasm localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 40x magnification in CellInsight CX7 LZR High-Content Screening (HCS) Platform (Product # CX7A1110LZR) and externally deconvoluted (D.Sage et al./Methods 115 (2017) 28–41.

Supportive validation

Supportive validation

PA3-023