Anti-CD34 antibody [EP373Y] - BSA and Azide free

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

This data was developed using the same antibody clone in a different buffer formulation (ab81289).

Flow cytometry overlay histogram showing left HUVEC positive cells and right negative HL60 stained with ab81289 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 mins. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab81289) (1x 10⁶ in 100μl at 0.2μg/ml (1/11550 dilution)) for 30 mins at 22°C.

The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed was incubated at 1/4000 dilution for 30 mins at 22°C

Isotype control antibody (black line) was Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human angiosarcoma labeling CD34 with unpurified ab81289 at 1/100-1/250.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Clone EP373Y (ab198395) has been successfully conjugated by Abcam. This image was generated using Anti-CD34 antibody [EP373Y] (Alexa Fluor^® 488). Please refer to ab195013 for protocol details.

Overlay histogram showing HUVEC cells stained with ab195013 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab195013, 1/500 dilution) for 30 min at 22°C.

Isotype control antibody (black line) was Rabbit IgG (monoclonal) Alexa Fluor^® 488 (ab199091) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.

This antibody gave a positive signal in HUVEC cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Triton X-100 for 15 min used under the same conditions.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human kidney tissue labelling CD34 with purified ab81289 at 1/2500. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as the secondary antibody at 1/500. Negative control using PBS instead of primary antibody. Counter stained with Hematoxylin.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Immunocytochemistry/Immunofluorescence analysis of HUVEC (Human umbilical vein endothelial cell line) cells labelling CD34 with purified ab81289 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (ab150077) (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counter stain.

Control : Secondary antibody only, ab150120, an Alexa Fluor^® 488-conjugated goat anti-mouse IgG (1/500).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Clone EP373Y (ab198395) has been successfully conjugated by Abcam. This image was generated using Anti-CD34 antibody [EP373Y] (PE). Please refer to ab223930 for protocol details.

Overlay histogram showing HUVEC cells stained with ab223930 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab223930, 1/500 dilution) for 30 min at 22°C.

Isotype control antibody (black line) was Rabbit IgG (monoclonal) Phycoerythrin (ab209478) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5,000 events were collected using a 50 mW Yellow/Green laser (561nm) and 586/15 bandpass filter.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Immunohistochemical analysis of formalin fixed paraffin embedded human kidney labelling CD34 with ab198395 at a concentration of 0.5µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with an OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was conducted for 32 min with ULTRA cell conditioning solution (CC1 pH 8.5). ab198395 anti CD34 antibody was incubated at 37°C for 16min. Sections were counterstained is with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

• IP

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Immunoprecipitation - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

CD34 was immunoprecipitated from TF-1 (Human bone marrow erythroleukemia cells) cells using purified ab81289 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab81289. Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated was used as secondary antibody at 1/1000 dilution. Blocking and dilution buffer and concentration : 5% NFDM/TBST

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

All lanes:

Immunoprecipitation - Anti-CD34 antibody [EP373Y] (<a href='/en-us/products/primary-antibodies/cd34-antibody-ep373y-ab81289'>ab81289</a>)

Predicted band size: 40 kDa

Observed band size: 120 kDa

false

• IHC-Fr

Lab

Immunohistochemistry (Frozen sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

This data was developed using ab81289, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat kidney tissue labeling CD34 with ab81289 at 1/50 (11.04 μg/mL) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). Positive staining on rat kidney. is observed. The nuclear counterstain was DAPI (Blue). Secondary antibody control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• IHC-P

AbReview54850****

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Paraformaldehyde-fixed, paraffin-embedded mouse kidney tissue stained for CD34 using ab81289 at 1/200 dilution in immunohistochemical analysis.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

This image is courtesy of an Abreview submitted by Rudolf Jung.

• IHC-P

AbReview63093****

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Paraformaldehyde-fixed, paraffin-embedded mouse prostate tissue stained for CD34 using ab81289 at 1/150 dilution in immunohistochemical analysis.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

This image is courtesy of an anonymous Abreview.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

IHC image of CD34 staining in Mouse normal brain formalin fixed paraffin embedded tissue section, performed on a Leica Bond^TM system using the standard protocol B. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with unpurified ab81289, 1/250 dilution, for 15 mins at room temperature. A Goat anti-Rabbit biotinylated secondary antibody was used to detect the primary, and visualized using an HRP conjugated ABC system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

• IHC-Fr

Lab

Immunohistochemistry (Frozen sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

This data was developed using ab81289, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat lung tissue labeling CD34 with ab81289 at 1/50 (11.04 μg/mL) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). Positive staining on rat lung. is observed. The nuclear counterstain was DAPI (Blue). Secondary antibody control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat kidney tissue labelling CD34 with purified ab81289 at 1/2500. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as the secondary antibody at 1/500. Negative control using PBS instead of primary antibody. Counter stained with Hematoxylin.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

• WB

Lab

Western blot - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

This data was developed using ab81289, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Negative control : Daudi, HL-60.

Low expression : HUVEC, Neuro-2a.

All lanes:

Western blot - Anti-CD34 antibody [EP373Y] (<a href='/en-us/products/primary-antibodies/cd34-antibody-ep373y-ab81289'>ab81289</a>) at 1/1000 dilution

Lane 1:

KG-1a (human bone marrow myeloblast) whole cell lysate at 20 µg

Lane 2:

TF-1 (human erythroleukemia erythroblast) whole cell lysate at 20 µg

Lane 3:

Daudi (human Burkitts lymphoma lymphoblast) whole cell lysate at 20 µg

Lane 4:

HL-60 (Human acute promyelocytic leukemia promyeloblast) whole cell lysate at 20 µg

Lane 5:

HUVEC (Human umbilical vein endothelial cell) whole cell lysate at 20 µg

Lane 6:

bEnd.3 (mouse brain endothelioma) whole cell lysate at 20 µg

Lane 7:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 8:

Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 80-120 kDa

false

Exposure time: 60s

• WB

Lab

Western blot - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

This data was developed using ab81289, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

This product failed to detect any signal in the kidney lysate.

All lanes:

Western blot - Anti-CD34 antibody [EP373Y] (<a href='/en-us/products/primary-antibodies/cd34-antibody-ep373y-ab81289'>ab81289</a>) at 1/1000 dilution

Lane 1:

Mouse lung tissue lysate at 20 µg

Lane 2:

Mouse kidney tissue lysate at 20 µg

Lane 3:

Mouse heart tissue lysate at 20 µg

Lane 4:

Rat lung tissue lysate at 20 µg

Lane 5:

Rat kidney tissue lysate at 20 µg

Lane 6:

Rat heart tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 80-120 kDa

false

Exposure time: 40s

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Normal uterus vessels tissue labeling CD34 with unpurified ab81289 at 1/100-1/250.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Normal kidney vessels tissue labeling CD34 with unpurified ab81289 at 1/100-1/250.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab81289).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

This IHC data was generated using the same anti-CD34 antibody clone, EP373Y, in a different buffer formulation (cat# ab81289).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Normal tonsil vessels tissue labeling CD34 with unpurified ab81289 at 1/100-1/250.

• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-CD34 antibody [EP373Y] - BSA and Azide free (AB198395)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.