Recombinant Anti-Synaptophysin antibody [YE269] (ab32127)

Lanes 1- 3: Merged signal (red and green). Green - ab32127 observed at 38 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.

ab32127 was shown to react with Synaptophysin in wild-type HEK-293T cells in Western blot. Loss of signal was observed when knockout sample ab257060 was used. Wild-type HEK-293T and SYP knockout whole cell lysates were subjected to SDS-PAGE. ab32127 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Immunocytochemistry/ Immunofluorescence analysis of mouse primary neuron cells labeling Synaptophysin with purified ab32127 at 1/100 (2.7µg/mL). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 µg/mL). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/mL) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
Confocal scanning Z step was set as 0.3 µm followed by image processing with maximum Z projection.

Formalin-fixed, paraffin-embedded sheep Gut (ileum) tissue stained for Synaptophysin using ab32127 at 1/10000 dilution in immunohistochemical analysis. The secondary antibody was a swein anti rabbit Biotin labelled, DAKO E0433 at 1/200 dilution.

Antigen Retrieval: Heat mediated - Buffer/Enzyme Used: CitricAcid buffer.

Blocking step: NGS/PBS as blocking agent for 30 minute(s). Concentration: 5%. Temperature: RT°C

Blocking/Dilution buffer: 5% NFDM/TBST

ab32127 staining synaptophysin in human iPS cell derived neurons by immunocytochemistry/immmunofluorescence.

Samples were fixed with paraformaldehyde and blocked with 1% serum for 30 minutes at room temperature. Samples were incubated with primary antibody at 1/250 dilution for 1 hour. ab150061 was used as the secondary antibody at 1/250 dilution. 

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Immunofluorescent staining of PC-12 (rat adrenal gland pheochromocytoma cell line) cells (fixed in 4% PFA, permeabilized with 0.1% Triton X 100) using purified ab32127 at a dilution of 1/50.

An Alexa Fluor^® 488 goat anti-rabbit antibody was used as the secondary at a dilution of 1/500 and the cells were counterstained with DAPI.

The negative control is shown in the bottom right hand panel - for the negative control, Alex Fluor^® 594 goat anti-mouse was used at a dilution of 1/500.

Immunohistochemical staining of paraffin embedded mouse cerebral cortex with purified ab32127 at a dilution of 1/400.

A pre-diluted HRP polymer for rabbit/mouse IgG was used as the secondary antibody and the sample was counterstained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

ab32127 staining Synaptophysin in rat brain tissue sections by Immunohistochemistry (IHC-Fr - frozen sections).

Tissue was fixed with formaldehyde, permeabilized with 0.2% Triton X-100 and blocked with 5% BSA for 2 hours at 25°C. Samples were incubated with primary antibody at 1/300 dilution for 10 hours at 4°C. An Alexa Fluor^® 555 rabbit polyclonal was used as a secondary antibody at 1/400 dilution.

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Immunohistochemical analysis of free floating Mouse striatum tissue stained for Synaptophysin (Green) using ab32127 at 1/500 dilution, followed by a AlexaFluor-488® donkey anti-rabbit secondary antibody at 1/10000 dilution. Image shows Expression of Synaptophysin in neuronal projections from cortex.

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Unpurified ab32127 showing positive staining in medulloblastoma tissue.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemical staining of paraffin embedded rat cerebral cortex with purified ab32127 at a dilution of 1/400.

A pre-diluted HRP polymer for rabbit/mouse IgG was used as the secondary antibody and the sample was counterstained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

Immunohistochemical staining of paraffin embedded human pancreas with purified ab32127 at a dilution of 1/400.

A pre-diluted HRP polymer for rabbit/mouse IgG was used as the secondary antibody and the sample was counterstained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

Unpurified ab32127 showing positive staining in lung neuroendocrine tumor tissue.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.