MAB305
Anti-Choline Acetyltransferase Antibody
CHEMICON®, mouse monoclonal, 1E6
Sinónimos:
ChAT, Choline Acetylase, CHOACTase
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Código UNSPSC:
12352203
eCl@ss:
32160702
NACRES:
NA.41
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Permítanos ayudarleNombre del producto
Anti-Choline Acetyltransferase Antibody, clone 1E6, ascites fluid, clone 1E6, Chemicon®
origen biológico
mouse
Nivel de calidad
forma del anticuerpo
ascites fluid
tipo de anticuerpo
primary antibodies
clon
1E6, monoclonal
reactividad de especies
human, rat, monkey
fabricante / nombre comercial
Chemicon®
técnicas
immunohistochemistry: suitable
isotipo
IgG1
Nº de acceso NCBI
Nº de acceso UniProt
Condiciones de envío
dry ice
modificación del objetivo postraduccional
unmodified
Información sobre el gen
human ... CHAT(1103)
rat ... Chat(290567)
rhesus monkey ... Chat(709977)
1 of 4
Este artículo | MAB5350 | AB15468 | AB143 |
|---|---|---|---|
| biological source mouse | biological source mouse | biological source chicken | biological source rabbit |
| species reactivity human, rat, monkey | species reactivity human, rat, guinea pig, monkey | species reactivity mouse | species reactivity mouse, human, feline, bat, rat, monkey |
| clone 1E6, monoclonal | clone 28C4, monoclonal | clone polyclonal | clone polyclonal |
| antibody form ascites fluid | antibody form purified immunoglobulin | antibody form affinity purified immunoglobulin | antibody form serum |
| Gene Information human ... CHAT(1103) | Gene Information human ... CHAT(1103) | Gene Information mouse ... Chat(12647) | Gene Information cat ... Chat(751512) |
| Quality Level 100 | Quality Level 100 | Quality Level 100 | Quality Level 100 |
Inmunógeno
Choline acetyltransferase purified from rat brain.
Aplicación
Detect Choline Acetyltransferase using this Anti-Choline Acetyltransferase Antibody, clone 1E6 validated for use in IH.
Immunohistochemistry: 1:100-1:250. See immunohistochmistry procedure below.
Optimal working dilutions must be determined by the end user.
IMMUNOHISTOCHEMISTRY PROCEDURE (PAP TECHNIQUE) FOR MAB305, MONOCLONAL ANTIBODY TO CHOLINE ACETYLTRANSFERASE
I) Perfusion & Sectioning Procedure
1. Perfuse through the heart with a fixative solution containing 4% paraformaldehyde in 0.12 M phosphate buffer (pH 7.3) for light microscopy (LM), and additionally, 0.1% gluteraldehyde and .002% CaCl2 for electron microscopy (EM).
2. Remove brain and postfix 2-18 hours at 4°C in 4% paraformaldehyde in 0.12 M phosphate buffer.
3. After brain is blocked for sectioning, wash in several changes of buffer for 2-3 hours.
4. Specimens for EM are sectioned on a Vibratome (50 μm) and rinsed in buffer, those for LM should be cryoprotected in 30% sucrose in buffer.
5. After freezing with dry ice, 30-40 μm thick sections of LM specimens are cut on a cryostat.
6. Sections are rinsed, and then stored in phosphate buffer containing 0.1% sodium azide.
II) Staining Procedure
Tissue is processed as freely-floating sections in continuously agitated solutions. All incubations are performed at room temperature unless otherwise stated.
1.a. For localizing ChAT-positive somata and dendrites:
Sections are washed in 0.1 M Tris-buffered saline (TBS; containing 1.4% NaCl, pH 7.3) only. No detergent or enzyme pretreatment is used.
b. For localizing ChAT-positive terminal-like structures:
Incubate sections in TBS (pH 8.1) for 5 minutes at 37°C. Transfer sections to TBS (pH 8.1) containing pronase (1.2 μg/mL) for 1 1/2-2 minutes at 37°C, followed by several ice cold buffer washes for a total of 5 minutes. The concentration of pronase and incubation time of the digestion should be evaluated for each region examined.
c. For localizing ChAT immunoreactivity and subsequently counterstaining the sections:
Incubation in TBS containing 0.1%-0.8% Triton X-100 for 15 minutes may increase the tissue penetration of the immunoreagents, but it also raises the background staining.
2. Incubate sections in normal goat serum (3-5%) for one hour. The working solutions of all antisera should also contain similarly diluted normal goat serum.
3. Incubate in anti-ChAT monoclonal antibody solution (Suggested working dilution 1:250, final working dilution must be determined by end user) for 2 hours at room temperature and then for an additional 6-18 hours at 4°C.
4. Incubate with second antibody (i.e. Goat anti-Mouse IgG, Cat. No.: AP124, dilution 1:50-100) for 1-2 hours.
5. Incubate with diluted PAP complex (i.e. Mouse PAP, Cat No.: PAP14, conc. 25-50 μg/mL) for one hour.
6. After rinsing in buffer, the second antibody and PAP steps are repeated for 40 minutes to 1 hour each in order to amplify staining intensity, particularly of small ChAT-containing structures.
7. React for 15 minutes with 0.06% 3,3′-diaminobenzidine×4 HCl (DAB; diluted in phosphate buffered saline, pH 7.3) and 0.006% H2O2.
8. Specimens for routine LM are postfixed for 1 minutes in 0.005% OsO4 (osmium tetraoxide), and then mounted, dehydrated and coverslipped. Selected regions blocked for EM are postfixed in 2% OsO4 for 1 hour, en bloc stained with uranyl acetate, and flat-embedded in Epon-Araldite resin.
Optimal working dilutions must be determined by the end user.
IMMUNOHISTOCHEMISTRY PROCEDURE (PAP TECHNIQUE) FOR MAB305, MONOCLONAL ANTIBODY TO CHOLINE ACETYLTRANSFERASE
I) Perfusion & Sectioning Procedure
1. Perfuse through the heart with a fixative solution containing 4% paraformaldehyde in 0.12 M phosphate buffer (pH 7.3) for light microscopy (LM), and additionally, 0.1% gluteraldehyde and .002% CaCl2 for electron microscopy (EM).
2. Remove brain and postfix 2-18 hours at 4°C in 4% paraformaldehyde in 0.12 M phosphate buffer.
3. After brain is blocked for sectioning, wash in several changes of buffer for 2-3 hours.
4. Specimens for EM are sectioned on a Vibratome (50 μm) and rinsed in buffer, those for LM should be cryoprotected in 30% sucrose in buffer.
5. After freezing with dry ice, 30-40 μm thick sections of LM specimens are cut on a cryostat.
6. Sections are rinsed, and then stored in phosphate buffer containing 0.1% sodium azide.
II) Staining Procedure
Tissue is processed as freely-floating sections in continuously agitated solutions. All incubations are performed at room temperature unless otherwise stated.
1.a. For localizing ChAT-positive somata and dendrites:
Sections are washed in 0.1 M Tris-buffered saline (TBS; containing 1.4% NaCl, pH 7.3) only. No detergent or enzyme pretreatment is used.
b. For localizing ChAT-positive terminal-like structures:
Incubate sections in TBS (pH 8.1) for 5 minutes at 37°C. Transfer sections to TBS (pH 8.1) containing pronase (1.2 μg/mL) for 1 1/2-2 minutes at 37°C, followed by several ice cold buffer washes for a total of 5 minutes. The concentration of pronase and incubation time of the digestion should be evaluated for each region examined.
c. For localizing ChAT immunoreactivity and subsequently counterstaining the sections:
Incubation in TBS containing 0.1%-0.8% Triton X-100 for 15 minutes may increase the tissue penetration of the immunoreagents, but it also raises the background staining.
2. Incubate sections in normal goat serum (3-5%) for one hour. The working solutions of all antisera should also contain similarly diluted normal goat serum.
3. Incubate in anti-ChAT monoclonal antibody solution (Suggested working dilution 1:250, final working dilution must be determined by end user) for 2 hours at room temperature and then for an additional 6-18 hours at 4°C.
4. Incubate with second antibody (i.e. Goat anti-Mouse IgG, Cat. No.: AP124, dilution 1:50-100) for 1-2 hours.
5. Incubate with diluted PAP complex (i.e. Mouse PAP, Cat No.: PAP14, conc. 25-50 μg/mL) for one hour.
6. After rinsing in buffer, the second antibody and PAP steps are repeated for 40 minutes to 1 hour each in order to amplify staining intensity, particularly of small ChAT-containing structures.
7. React for 15 minutes with 0.06% 3,3′-diaminobenzidine×4 HCl (DAB; diluted in phosphate buffered saline, pH 7.3) and 0.006% H2O2.
8. Specimens for routine LM are postfixed for 1 minutes in 0.005% OsO4 (osmium tetraoxide), and then mounted, dehydrated and coverslipped. Selected regions blocked for EM are postfixed in 2% OsO4 for 1 hour, en bloc stained with uranyl acetate, and flat-embedded in Epon-Araldite resin.
Research Category
Neuroscience
Neuroscience
Research Sub Category
Neurotransmitters & Receptors
Neuronal & Glial Markers
Neurotransmitters & Receptors
Neuronal & Glial Markers
Acciones bioquímicas o fisiológicas
Recognizes cholinergic neurons in the brain and spinal cord (CNS).
Forma física
Ascites fluid containing no preservatives.
Unpurified
Nota de preparación
Maintain for 1 year at -20°C from date of shipment. Aliquot to avoid repeated freezing and thawing. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
Nota de análisis
Control
Brain tissue
Brain tissue
Otras notas
Concentration: Please refer to the Certificate of Analysis for the lot-specific concentration.
Información legal
CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany
Cláusula de descargo de responsabilidad
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
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Código de clase de almacenamiento
10 - Combustible liquids
Clase de riesgo para el agua (WGK)
WGK 1
Punto de inflamabilidad (°F)
Not applicable
Punto de inflamabilidad (°C)
Not applicable
Certificados de análisis (COA)
Busque Certificados de análisis (COA) introduciendo el número de lote del producto. Los números de lote se encuentran en la etiqueta del producto después de las palabras «Lot» o «Batch»
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