DETAILED DESCRIPTION OF THE INVENTION The present invention relates to methods and reagents for determining ligands in biological fluids such as serum, plasma, spinal fluid, amnionic fluid and urine.
The term ligand as used in the present invention refers to haptens, polypeptides, proteins, and glycoproteins having molecular weights generally below 150,000.
Haptens are protein-free bodies, generally of low molecular weight that do not induce antibody formation when injected into an animal, but are reactive to antibodies.
Antibodies to hapten are raised by first conjugating the hapten to a protein and injecting the conjugate product into an animal or human.
The resulting antibodies are isolated by conventional antibody isolation techniques.
For purposes of the present invention, the antibodies should be substantially free of serum protein materials such as indicator enzymes used in the test or inhibitors to antibody binding.
These are conveniently removed by ion exchange chromatography on an anion exchange column or other suitable protein separation technique.
Representative ligands determinable by methods of the present invention are steroids such as estrone, estradiol, cortisol, testosterone, progesterone, chenodeoxycholic acid, digoxin, cholic acid, deoxycholic acid, lithocholic acids and the ester and amide derivatives thereof; vitamins such as vitamin B-12, folic acid; thyroxine, triiodothyronine, histamine, serotonin, prostaglandins such as PGE, PGF, PGA; adrenalin, noradrenalin and drugs such as opiates, theophylline, dilantin; barbituates such as phenobarbitol and derivatives thereof and carbamazepins, aminoglycoside antibiotics like gentimycin and tobramycin.
Representative polypeptides and glycoproteins determinable by methods of the present invention are insulin, platelet factor 4 and polypeptide determinants of large antigens.
Ligand analogs are functional or functionalized ligands suitable for conjugation to irreversible enzyme inhibitors.
Acids, esters, amides, amines, hydroxy, isocyanate, isothiocyanate are suitable functional groups
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Molecular Biology 
