Anti-Tau antibody [E178] (ab32057) at 1/100 dilution (Purified) + Human brain lysates at 15 µg

Secondary
Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 79 kDa
Observed band size: 55-74 kDa why is the actual band size different from the predicted?

The observed molecular weights are consistent with what have been described in literature PMID: 9276470

Immunohistochemistry (Frozen sections) analysis of rat cerebrum tissue sections labeling Tau with purified ab32057 at 1/500 (0.2 µg/ml). Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. DAPI was used as a counterstain.

IHC image of Tau staining in a section of frozen normal human Alzheimer brain performed on a Leica BOND^TM system using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab32057, 1/1000 dilution, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.

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.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human cerebrum tissue sections labeling Tau with purified ab32057 at 1/4000 dilution (0.03 µg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0) before commencing with IHC staining protocol. ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

ab32057 (purified) at 1/20 dilution (0.5ug) immunoprecipitating Tau in Human fetal brain lysates.
Lane 1: Human fetal brain lysates 10ug
Lane 2 (+): ab32057 & Human fetal brain lysates
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32057 in Human fetal brain lysates
For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
Blocking and diluting buffer: 5% NFDM/TBST.

Anti-Tau antibody [E178] (ab32057) at 1/5000 dilution (Purified) + Mouse brain lysates at 15 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 79 kDa
Observed band size: 55-74 kDa why is the actual band size different from the predicted?

The observed molecular weights are consistent with what have been described in literature PMID: 9276470

Immunohistochemistry (Frozen sections) analysis of mouse cerebrum tissue sections labeling Tau with purified ab32057 at 1/500 (0.2 µg/ml). Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. DAPI was used as a counterstain.

All lanes : Anti-Tau antibody [E178] (ab32057) at 1/1000 dilution

Lane 1 : Untreated SH-SY5Y cell
lysate
Lane 2 : SH-SY5Y cell
lysate SH SY5Y cell lysate treated with Oka/CalA. Cells are serum-starved overnight, and then treated with 1nM calyculin A and 500nM Okadaic acid for 2 hours at 37&degC.

Predicted band size: 79 kDa

Immunohistochemistical detection of Tau antibody [E178] (ab32057) on formaldehyde fixed paraffin-embedded rat tongue sectionsAntigen retrieval step: Heat mediated. Buffer Used: Citric acid pH6. Permeabilization: None. Primary antibody incubated at 1/1000 for 2 hours @ 21°C in TBS/BSA/azide. Secondary antibody: anti Rabbit IgG Conjugated to Biotin (1/200). A strong pattern of immunostaining which appears to be mostly localised to nerve fibres and their cell bodies (Islet of Langerhans cells are also positive). In submitted image of central area of tongue coloured arrowheads indicate features: red for nerves cut in cross-section (T/S), each brown dot representing a single axon green for what appears to me to be small nerve fibres wrapping around a partial muscle fibre black for a Ganglion containing seven positive nerve cell bodies. Surrounding these are collagen fibres (C), adipocytes (A) and skeletal/striated muscle fibres in L/S ( M-

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Immunohistochemistical detection of Tau antibody [E178] (ab32057) on formaldehyde-fixed paraffin-embedded human salivary gland sections. Antigen retrieval step: Heat mediated.  Buffer Used: Citric acid pH6. Permeabilization: No.  Blocking step: 1% BSA for 10 mins @ 21°C. ab32057 incubated at 1/1000 for 2 hours @  21°C in TBS/BSA/azide. Secondary antibody:  anti rabbit IgG conjugated to Biotin (1/200). NB: An interesting pattern of positivity that seems to be supported by the Human Protein Atlas. Coloured arrowheads in the submitted image indicate features: red for positive serous glands, blue for positive intra-lobular collecting ducts, black for negative mucous glands (there is a serous demilune around this acinus), yellow for intralobular collecting ducts, green for nerve tracks in the interlobular areas, blue for positive interlobular collecting ducts. There appears to be a population of positive nuclei but this may b

Ab32057, at a dilution of 1/500, staining Tau in paraffin embedded human brain sections by Immunohistochemistry. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.