Technical Data
N2700-03
Nitrotyrosine (nTyr)
Description:
Nitrotyrosine is formed in tissues in presence of the active metabolite NO and is a stable end product of nitrosylation of tyrosine. Inflammation is characterized by increased nitric oxide (NO) production. NO reacts rapidly with superoxide to form peroxynitrite. At physiological pH and in the presence of transition metals, peroxynitrite undergoes heterolytic cleavage to form hydroxyl anion and nitronium ion, the latter of which nitrates protein tyrosine residues. The presence of nitrotyrosine has been detected in various inflammatory processes including atherosclerotic plaques, Amyotrophic Lateral Sclerosis (ALS) and Multiple Sclerosis (MS). Thus, the presence of nitrotyrosine on proteins can be used as a marker for peroxynitrite formation in vivo and consequently as a marker of NO-mediated tissue damage.

Applications:
Suitable for use in ELISA, Western Blot and Immunohistochemistry. Other applications have not been tested.

Recommended Dilutions:
Western Blot: 1:50 for reduced and no-reduced samples. Block with 5% BSA or skimmed milk.
Immunohistochemistry (Frozen, cytospins): 1:50 with acetone fixation for 10 minutes at -20 C. Block endogenous peroxidase with 0.3% H2O2 in PBS (or methanol for intracellular staining). Block with 10% NGS or 5% BSA for 30 minutes.
Immunohistochemistry (Paraffin): 1:200-1:400 with 10% formalin fixation, Use 3% H2O2 to block endogenous peroxidases. Use citrate buffer pH 6.0 for 1 minute at 100C as antigen retrieval treatment.
Optimal dilutions to be determined by the researcher.

Western Blot Controls:
Mouse kidney lysate, mouse optic nerve, retina , spinal cord and brain lysates, rat aorta lysate

Storage and Stability:
May be stored at 4C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20C. Aliquots are stable for at least 12 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
TypeIsotypeCloneGrade
MabIgG2b6A610Affinity Purified
SizeStorageShippingSourceHost
100ug-20CBlue IceMouse
Concentration:
~0.1mg/ml
Immunogen:
Nitrated KLH
Purity:
Purified by Protein G affinity chromatography.
Form
Supplied as a liquid in PBS, 0.1% BSA, 0.02% sodium azide.
Specificity:
Recognizes nitrotyrosine, both with the free amino acid as well as with proteins containing nitrotyrosine. Does not crossreact with Phosphotyrosine or Chlorotyrosine.
Intended for research use only. Not for use in human, therapeutic, or diagnostic applications.
1. ter Steege, JCA et al; Nitrotyrosine in plasma of celiac disease patients as detected by a new sandwich ELISA. Free Radic Biol Med 1998, 25:953. 2. Beckman, JS et al; Extensive nitration of protein tyrosines in human atherosclerosis detected by immunohistochemistry. Biol Chem 1994, 375:81. 3. Kooy, NW et al; Evidence for in vivo peroxynitrite production in human acute lung injury. Am J Respir Crit Care Med 1995, 151:1250. 4. Beckman, JS et al; Apperent hydroxyl radical production by peroxynitrite: Implications of endothelial injury from nitric oxide and superoxide. Proc Natl Acad Sci USA 1990, 87:1620.