MA5-56385

antibody from Invitrogen Antibodies

Targeting: SPON1 f-spondin, KIAA0762

Antibody data

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Validation data

Product number: MA5-56385 - Provider product page
Provider: Invitrogen Antibodies
Product name: SPON1 Recombinant Rabbit Monoclonal Antibody (PSH02-84)
Antibody type: Monoclonal
Antigen: Synthetic peptide
Description: Positive Control: SW1990 cell lysate, PANC-1 cell lysate, U-2 OS cell lysate, HepG2 cell lysate, PANC-1, SW1990. Tissue Specificity: Low tissue specificity. Subcellular Location: Secreted protein; extracellular space; extracellular matrix. Sequence Similarities: 100% Mouse/Rat. Predicted band size: 91 kDa.
Reactivity: Human
Host: Rabbit
Isotype: IgG
Antibody clone number: PSH02-84
Vial size: 100 µL
Concentration: 1 mg/mL
Storage: Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

SPON1 protein structure - MA5-56385 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using SPON1 Recombinant Rabbit Monoclonal Antibody (PSH02-84) (Product # MA5-56385) and 90 kDa bands corresponding to SPON1 was observed across cell lines tested except HEL 92.1.7. Whole cell extracts (30 µg lysate) of NIH:OVCAR-3 (Lane 1), PANC-1 (Lane 2), U2-OS (Lane 3), Caki-1 (Lane 4), SK-N-MC (Lane 5) and HEL 92.1.7 (Lane 6) 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™ 3 Western Blot Transfer Device (Product # IB31001). The blot was probed with the primary antibody (1:2,000 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 Pico PLUS Chemiluminescent Substrate (Product # 34580).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using SPON1 Recombinant Rabbit Monoclonal Antibody (PSH02-84) (Product # MA5-56385) and 91-120 kDa bands corresponding to F-spondin was observed across cell lines tested. Whole cells extracts (20 µg lysate) of SW1990 (Lane 1), PANC-1 (Lane 2), U-2 OS (Lane 3) and HepG2 (Lane 4) were electrophoresed using 4-20% SDS-PAGE gel. Resolved proteins were transferred onto a PVDF membrane. The blot was blocked with 5% NFDM/TBST for 1 hour at room temperature, then probed with the primary antibody (1:1,000 dilution) at 4 degrees Celsius overnight and detected by chemiluminescence with HRP labeled Goat anti-Rabbit IgG secondary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of F-spondin using SW1990 cells. The cells were fixed 100% pre-chilled methanol for 5 minutes, and blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. The cells labeled with SPON1 Recombinant Rabbit Monoclonal Antibody (PSH02-84) (Product # MA5-56385) (green) at 1:100 dilution and beta tublin (red) at 1:100 dilution in 1% BSA in PBST overnight at 4 degrees Celsius and then with iFluor™ 488 Goat Anti-Rabbit IgG H&L and iFluor™ 594 Goat Anti-Mouse IgG H&L secondary antibodies at 1:1,000 dilution for 1 hour at room temperature. Nuclei were stained with DAPI (blue). The images were captured at 200X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of F-spondin using PANC-1 cells. The cells were fixed 4% paraformaldehyde for 20 minutes, permeabilized with 0.1% Triton™ X-100 in PBS for 5 minutes, and blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. The cells labeled with or without SPON1 Recombinant Rabbit Monoclonal Antibody (PSH02-84) (Product # MA5-56385) (green) at 1:500 dilution in 1% BSA in PBST overnight at 4 degrees Celsius and then with iFluor™ 488 Goat Anti-Rabbit IgG H&L secondary antibody at 1:1,000 dilution for 1 hour at room temperature. Nuclei were stained with DAPI (blue). The images were captured at 200X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of SPON1 was performed using 70% confluent log phase PANC-1 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with SPON1 Recombinant Rabbit Monoclonal Antibody (PSH02-84) (Product # MA5-56385) 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), (1:2,000 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 dilution). Panel d represents the merged image showing golgi like localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 40X magnification.