PMN Rabbit Polyclonal Antibody

CAT#: AP31661SU-N

PMN rabbit polyclonal antibody, Serum



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CNY 6,468.00


货期*
5周

规格
    • 2 ml

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Specifications

Product Data
Applications CT, IHC
Recommend Dilution Cytotoxic assasy.
Immunohistochemistry on frozen and paraffin sections.

For Cytotoxic Antibodies:
Modified Colorimetric Microtiter Assay 1
Results: Antisera of the above this antibody diluted 1/20 exhibits >80% cytotoxicity on rat PMNs
Antisera diluted 1/20 exhibits
<15% cytotoxicity on rat thymocytes or splenocytes.
For Agglutinating Antibodies:
Antisera dilutions in RPMI-1640 incubated with target cells at 4°C-8°C for 1hr.
Agglutination determined by microscopic observations.
Results: Antisera of the above this antibody strongly
agglutinates rat PMNs but not lymphocytes at dilutions 1/20 to 1/200
Reactivity Rat
Host Rabbit
Clonality Polyclonal
Specificity This antibody reacts to PMN.
Formulation State: Serum
State: Liquid serum
Conjugation Unconjugated
Storage Condition Upon receipt, store (in aliquots) at -20°C to -80°C.
Avoid repeated freezing and thawing.
Background CLAD51140 is an antiserum directed against rat PMNs (Polymorphonuclear
Leukocytes). This antisera is adsorbed repeatedly with red blood cells and lymphocytes
to remove most antibodies to antigens common among different cell types. This
adsorption significantly improves cytotoxic specificity.
Synonyms Polymorphonuclear Leukocytes
Note Protocol: Rabbit anti-Rat PMN IHC protocol
(can also be used for mouse sections)
Sections:
Animals were perfused with 10 mM PBS, pH 7.4 followed by 4% paraformaldehyde in 100 mM phosphate buffer, pH 7.4. Tissue was removed and placed in 4% paraformaldehyde for 6 hours at room temperature, then transferred to 10 mM PBS, pH 7.4 in 0.15M isotonic saline at 4°C overnight. Sections were cut at 50 μm on a vibratome using 10 mM PBS and placed in tissue culture wells at 4°C overnight.
Procedure:
1. Pretreat slices in 50 mM NH4Cl (0.267g/100 ml PBS) in PBS at 22°C for 1 hour.
2. Pretreat slices in 0.1% Triton X-100 in PBS at 22°C for 1 hour.
3. Wash in PBS for 5 minutes at 22°C.
4. Block in PBS with 5% NGS at 22°C for 2 hours.
5. Incubate in primary antibody (Rabbit anti-rat PMN), AP31661SU-N, diluted 1/3000 in PBS with 5% NGS overnight at 4°C.
6. Wash in PBS with 5% NGS at 22°C for 30 minutes.
7. Incubate in secondary antibody, biotinylated goat anti-rabbit IgG in PBS with 5% NGS at 22°C for 1 hour, diluted:
1) 1/200, 125 μl/25 ml
2) Per instructions, Vector ABC Elite Kit, 8 drops/25 ml
8. Wash in PBS with 5% NGS at 22°C for 30 minutes.
9. Block endogenous peroxidase activity. Immediately before use, mix 81 ml PBS, 9 ml methanol and 10 ml 30% H2O2. Incubate for 10 minutes at 22°C.
10. Wash in PBS only 1 x 10 minutes at 22°C.
11. Wash in PBS only 1 x 20 minutes at 22°C
12. Wash in PBS with 5% NGS at 22°C for 30 minutes.
Prepare ABC reagent, if using this option.
13. Incubate in:
1) KPL Streptavidin-peroxidase conjugate diluted 1/200 (125 μl/25 ml) in PBS with 5% NGS, 0.1% Tween 20.
2) Vector ABC elite, diluted according to kit instructions for 1 hour at 22°C.
14. Wash in PBS with 5% NGS at 22°C for 10 minutes.
15. Incubate with DAB (2-5 minutes). Stop reaction with PBS wash.
16. Allow to air dry.
17. Wash salts off in ddH2O. Dehydrate, clear and mount on chromalum gelatin coated slides.
Notes:
Paraformaldehyde:
2.76g monobasic
21.45g dibasic heptahydrate
1000 ml ddH2O
40g Paraformaldehyde
PBS (10mM):
0.276 monobasic
2.15g dibasic heptahydrate
100 ml ddH2O
8.76 NaCl/1000 ml
900 ml Isotonic saline + 100 ml mono/dibasic
pH to 7.4
Primary antibody: 17 μl/50 ml PBS with 5% NGS
Reference Data
*Delivery time may vary from web posted schedule. Occasional delays may occur due to unforeseen complexities in the preparation of your product. International customers may expect an additional 1-2 weeks in shipping.
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