Detection Limit
The lowest concentration of mouse NGAL giving an absorbance reading greater than 2 SD above the mean zero (calibrator 1) reading was 0.75 pg/ mL, which is significantly lower than the value of the lowest calibrator concentration above zero (10 pg/mL).
General Description
NGAL (neutrophil gelatinase-associated lipocalin) belongs to the lipocalin family of proteins. These are secreted proteins characterized by 1) their ability to bind small hydrophobic molecules in a structur-ally conserved pocket formed by β-pleated sheet, 2)to bind to specific cell-surface receptors, and 3) to form macromolecular complexes. NGAL has many synonyms: perhaps the most widely used is lipocalin 2 (LCN 2); more recently the name siderocalin has been used to express NGAL's ability to bind bacterial siderophores. In the mouse, NGAL was first known at the mRNA level as 24p3 and was subsequently also investigated under the names 24-kDa superinducible protein (SIP24) or uterocalin.
Mouse NGAL consists of a single disulfidebridged polypeptide chain of 180 amino-acid residues with a calculated molecular mass of 20.9 kDa, but glycosylation increases its apparent molecular mass on SDS-PAGE to 24 kDa. In some situations mouse NGAL may be co-expressed and form complexes with matrix metalloproteinase-9 (MMP-9), but how this complex is linked is unknown, given that mouse NGAL, unlike its human homologue, does not possess a third cysteinyl residue capable of forming an intermolecu-lar disulfide bridge.
In the adult mouse, NGAL is expressed in neutrophil polymorphonuclear leucocytes, luminal and glandular epithelial cells of the uterus, and in the lung, spleen, vagina and epididymis. It may be expressed in additional cell types during embryonic develop-ment and in response to various stimuli. This applies to certain kidney cells (see below), LPS-stimulated macrophages, dexamethasone-stimulated L-cells, and basic fibroblast growth factor-stimulated fibroblasts. Moreover, mouse NGAL is a type-1 acute-phase protein, being secreted by hepatocytes in the in-vivo turpentine model or in response to TNF-a or dexamethasone in vitro.
NGAL and acute kidney injury
Apart from the expression of mouse NGAL that occurs in the above situations, NGAL undergoes an early and dramatic upregulation in mouse kidney cells after infection with SV40 or polyoma virus, and this was in fact the ffrst indication of the existence of NGAL and its early response in injured kidney cells.
Years later this ffnding was extended to postischemic and nephrotoxic injury. The marked upregulation of NGAL mRNA and protein levels in the early post-ischemic mouse kidney was detected predominantly in proximal tubule cells and NGAL was easily detected by Western blotting of urine after ischemia and cisplatin-induced nephrotoxicity.
Urinary NGAL has also been found to be raised in mouse models of diabetic and obstructive nephropathy. However, there is a paucity of quantitative data on NGAL levels in mouse urine, plasma or serum, whether in the basal state or after kidney injury. Most analyses have been by Western blotting, which has not been able to detect NGAL in serum except after induction by an acute phase reaction.
It nevertheless appears that the determination of urinary NGAL may become a convenient end-point in mouse models of renal injury, nephrotoxicity and other nephropathies.
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