Following is a data sheet to one of our NOS antibodies which includes some
good references to help get you started. If you have any questions, get
back to me. Good Luck.
Regards,
James Stiehr
Affinity BioReagents, Inc.
jcim at pipeline.com
908-369-3799
Product Data:
Catalog Number Size
PA3-030 100 µl
Description: Polyclonal (Rabbit) Anti-Inducible Nitric Oxide Synthase
(iNOS) Antibody
Specificity: Nitric Oxide (NO) is an inorganic, gaseous free radical that
carries a variety of messages between cells including vasorelaxation,
neurotransmission and cytotoxicity. NO is produced by a Nitric Oxide
Synthase (NOS) family of enzymes which oxidizes the amino acid L-arginine
to form L-citrulline and NO. Two constitutive forms, brain NOS (bNOS) &
endothelial cell NOS (eNOS), and one inducible form (iNOS), have been
cloned showing regions for binding of calmodulin, NADPH, FAD, and FMN.
iNOS has been found in a variety of tissues including macrophages,
hepatocytes, synoviocytes, and smooth muscle cells. It is induced after
exposure to cytokines such as interferon-* (IFN), tumor necrosis factor,
interleukin-1, interleukin-2, and lipopolysaccarides (LPS). Studies show
that iNOS contains a tightly bound calmodulin, and acts independently of
calcium levels, unlike eNOS and bNOS which are Ca2+/calmodulin dependent.
After cytokine induction, iNOS has a delayed response followed by high
levels of NO produced over a long period of time. New Zealand White Rabbits
were immunized with a synthetic peptide with a sequence derived from the
C-terminal end of mouse macrophage NOS, amino acids 1131-1144, with one
additional N-terminal cysteine residue added for conjugation to KLH.
Peptide Sequence: C - K1131 - K - G - S - A - L - E - E - P - K - A - T - R
- L1144
Uses*: This polyclonal antibody has been successfully used to detect iNOS
in Western blot and immunohistochemical experiments in samples first
induced with cytokines. It shows no cross reactivity with eNOS or bNOS.
By Western blot, PA3-030 detects an approximately 130 kDa band in human,
rat and mouse samples as shown in chart. In immunocytochemical
experiments, diffuse staining is seen after induction with both IFN and
LPS, but is weaker after induction by only one.
SuggestedWorkingDilution: Various assay conditions require that the optimal
working concentrations be determined by serial dilution of this product.
Suggested starting dilution is: Western blot: 1: 2000
Immunocytochemistry: 1: 500
Form & Storage: 100 µl of polyclonal antiserum diluted in PBS. Store at
-20oC. Avoid repeated freeze/thaw cycles. Contains Sodium Azide.
References: P. Feldman, et al. The Surprising Life of Nitric Oxide.
Chemical & Engineering News, 71(51): 26-38, 1993.M. Stefanovic-Racic, et
al. Nitric Oxide and Arthritis. Arthritis & Rheumatism, 36(8): 1036-1044,
1993.C. Nathan. Nitric Oxide as a Secretory Product of Mammalian Cells.
The FASEB Journal, 6: 3051-3064, 1992.C. Lyons, G. Orloff & J. Cunningham.
Molecular Cloning and Functional Expression of an Inducible Nitric Oxide
Synthase from a Murine Macrophage Cell Line. J. Biol. Chem. 267(9):
6270-6374, 1992.C. Lowenstein, et. al. Cloned and Expressed Macrophage
Nitric Oxide Synthase Contrasts with the Brain Enzyme. Proc. Natl. Acad.
Sci. 89: 6711-6715, 1992.D. Bredt, et. al. Cloned and Expressed Nitric
Oxide Synthase Structurally Resembles Cytochrome P-450 Reductase. Nature,
351: 714-718, 1991.
*This Product is for Research Use Only and is Not Intended for Use in
Humans or Clinical Diagnosis.
