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2.
Mol Pharmacol ; 60(6): 1399-406, 2001 Dec.
Article in English | MEDLINE | ID: mdl-11723248

ABSTRACT

It was previously shown that CYP3A4 is induced in the human intestinal Caco-2 cell model by treatment with 1alpha,25-dihydroxy vitamin D3 (1,25-D3). We demonstrate the vitamin D analog, 19-nor-1alpha,25-dihydroxy vitamin D2, is also an effective inducer of CYP3A4 in Caco-2 cells, but with half the potency of 1,25-D3. We report that treatment of LS180 cells, a human intestinal cell line, with 1 to 10 nM 1,25-D3 dose dependently increased CYP3A4 protein and CYP3A4 mRNA expression. CYP3A4- and CYP3A23-promoter-Luciferase reporter constructs transiently transfected into LS180 cells were transcriptionally activated in a dose-dependent manner by 1,25-D3, whereas mutation of the nuclear hormone receptor binding motif (ER6) in the CYP3A4 promoter abrogated 1,25-D3 activation of CYP3A4. Although the CYP3A4 ER6 promoter element has been shown to bind the pregnane X receptor (PXR), this receptor does not mediate 1,25-D3 induction of CYP3A4 because a) PXR is not expressed in Caco-2 cells; b) PXR mRNA expression is not induced by 1,25-D3 treatment of LS180 cells; and c) the ligand binding domain of human PXR was not activated by 1,25-D3. 1,25-D3 uses the vitamin D receptor to induce CYP3A4 because a) the vitamin D receptor (VDR)-retinoid X receptor (RXR) heterodimer binds specifically to the CYP3A4 ER6; b) selective mutation of the CYP3A4 ER6 disrupted the binding of VDR-RXR; and c) reporter constructs containing only three copies of the CYP3A4 ER6 linked to a TK-CAT reporter were activated by 1,25-D3 only in cells cotransfected with a human VDR expression plasmid. These data support the hypothesis that 1,25-D3 and VDR induce expression of intestinal CYP3A by binding of the activated VDR-RXR heterodimer to the CYP3A PXR response element and promoting gene transcription.


Subject(s)
Calcitriol/pharmacology , Cytochrome P-450 Enzyme System/biosynthesis , Intestines/drug effects , Mixed Function Oxygenases/biosynthesis , Transcription, Genetic/drug effects , Animals , COS Cells , Caco-2 Cells , Cells, Cultured , Cytochrome P-450 CYP3A , Cytochrome P-450 Enzyme System/genetics , Dimerization , Dose-Response Relationship, Drug , Enzyme Induction , Ergocalciferols/pharmacology , Haplorhini , Humans , Intestines/enzymology , Luciferases/genetics , Mixed Function Oxygenases/genetics , Pregnane X Receptor , Protein Binding , Receptors, Calcitriol/genetics , Receptors, Calcitriol/metabolism , Receptors, Cytoplasmic and Nuclear/metabolism , Receptors, Retinoic Acid/metabolism , Receptors, Steroid/metabolism , Retinoid X Receptors , Time Factors , Transcription Factors/metabolism , Transcriptional Activation
3.
Pharmacogenetics ; 11(7): 555-72, 2001 Oct.
Article in English | MEDLINE | ID: mdl-11668216

ABSTRACT

The pregnane X receptor (PXR)/steroid and xenobiotic receptor (SXR) transcriptionally activates cytochrome P4503A4 (CYP3A4) when ligand activated by endobiotics and xenobiotics. We cloned the human PXR gene and analysed the sequence in DNAs of individuals whose CYP3A phenotype was known. The PXR gene spans 35 kb, contains nine exons, and mapped to chromosome 13q11-13. Thirty-eight single nucleotide polymorphisms (SNPs) were identified including six SNPs in the coding region. Three of the coding SNPs are non-synonymous creating new PXR alleles [PXR*2, P27S (79C to T); PXR*3, G36R (106G to A); and PXR*4, R122Q (4321G to A)]. The frequency of PXR*2 was 0.20 in African Americans and was never found in Caucasians. Hepatic expression of CYP3A4 protein was not significantly different between African Americans homozygous for PXR*1 compared to those with one PXR*2 allele. PXR*4 was a rare variant found in only one Caucasian person. Homology modelling suggested that R122Q, (PXR*4) is a direct DNA contact site variation in the third alpha-helix in the DNA binding domain. Compared with PXR*1, and variants PXR*2 and PXR*3, only the variant PXR*4 protein had significantly decreased affinity for the PXR binding sequence in electromobility shift assays and attenuated ligand activation of the CYP3A4 reporter plasmids in transient transfection assays. However, the person heterozygous for PXR*4 is normal for CYP3A4 metabolism phenotype. The relevance of each of the 38 PXR SNPs identified in DNA of individuals whose CYP3A basal and rifampin-inducible CYP3A4 expression was determined in vivo and/or in vitro was demonstrated by univariate statistical analysis. Because ligand activation of PXR and upregulation of a system of drug detoxification genes are major determinants of drug interactions, it will now be useful to extend this work to determine the association of these common PXR SNPs to human variation in induction of other drug detoxification gene targets.


Subject(s)
Alleles , Aryl Hydrocarbon Hydroxylases , Receptors, Cytoplasmic and Nuclear/chemistry , Receptors, Cytoplasmic and Nuclear/genetics , Receptors, Steroid/chemistry , Receptors, Steroid/genetics , Xenobiotics/metabolism , Amino Acid Sequence , Animals , Chromosome Mapping/methods , Cytochrome P-450 CYP3A , Cytochrome P-450 Enzyme System/genetics , Cytochrome P-450 Enzyme System/metabolism , Humans , Models, Molecular , Molecular Sequence Data , Oxidoreductases, N-Demethylating/genetics , Oxidoreductases, N-Demethylating/metabolism , Polymorphism, Single Nucleotide/genetics , Pregnane X Receptor , Receptors, Cytoplasmic and Nuclear/physiology , Receptors, Steroid/physiology , Sequence Homology, Amino Acid , Transcriptional Activation/physiology
4.
J Biol Chem ; 276(42): 39411-8, 2001 Oct 19.
Article in English | MEDLINE | ID: mdl-11509573

ABSTRACT

Sister of P-glycoprotein (SPGP) is the major hepatic bile salt export pump (BSEP). BSEP/SPGP expression varies dramatically among human livers. The potency and hierarchy of bile acids as ligands for the farnesyl/bile acid receptor (FXR/BAR) paralleled their ability to induce BSEP in human hepatocyte cultures. FXR:RXR heterodimers bound to IR1 elements and enhanced bile acid transcriptional activation of the mouse and human BSEP/SPGP promoters. In FXR/BAR nullizygous mice, which have dramatically reduced BSEP/SPGP levels, hepatic CYP3A11 and CYP2B10 were strongly but unexpectedly induced. Notably, the rank order of bile acids as CYP3A4 inducers and activators of pregnane X receptor/steroid and xenobiotic receptor (PXR/SXR) closely paralleled each other but was markedly different from their hierarchy and potency as inducers of BSEP in human hepatocytes. Moreover, the hepatoprotective bile acid ursodeoxycholic acid, which reverses hydrophobic bile acid hepatotoxicity, activates PXR and efficaciously induces CYP3A4 (a bile-metabolizing enzyme) in primary human hepatocytes thus providing one mechanism for its hepatoprotection. Because serum and urinary bile acids increased in FXR/BAR -/- mice, we evaluated hepatic transporters for compensatory changes that might circumvent the profound decrease in BSEP/SPGP. We found weak MRP3 up-regulation. In contrast, MRP4 was substantially increased in the FXR/BAR nullizygous mice and was further elevated by cholic acid. Thus, enhanced hepatocellular concentrations of bile acids, due to the down-regulation of BSEP/SPGP-mediated efflux in FXR nullizygous mice, result in an alternate but apparent compensatory up-regulation of CYP3A, CYP2B, and some ABC transporters that is consistent with activation of PXR/SXR by bile acids.


Subject(s)
ATP-Binding Cassette Transporters/biosynthesis , Cell Nucleus/metabolism , Cytochrome P-450 Enzyme System/chemistry , Cytochrome P-450 Enzyme System/metabolism , ATP Binding Cassette Transporter, Subfamily B, Member 11 , ATP-Binding Cassette Transporters/genetics , Amino Acid Sequence , Animals , Base Sequence , Bile Acids and Salts/metabolism , Cell Line , Cells, Cultured , Cytochrome P-450 CYP3A , Dimerization , Dose-Response Relationship, Drug , Down-Regulation , Genes, Reporter , Hepatocytes/metabolism , Humans , Immunoblotting , Ligands , Liver/metabolism , Luciferases/metabolism , Mice , Mice, Inbred BALB C , Mice, Transgenic , Mixed Function Oxygenases/metabolism , Molecular Sequence Data , Multidrug Resistance-Associated Proteins/metabolism , Plasmids/metabolism , Promoter Regions, Genetic , Protein Binding , Protein Structure, Tertiary , Reverse Transcriptase Polymerase Chain Reaction , Ribosomal Proteins/metabolism , Sequence Homology, Nucleic Acid , Transfection , Up-Regulation , Ursodeoxycholic Acid/pharmacology
5.
Nat Genet ; 27(4): 383-91, 2001 Apr.
Article in English | MEDLINE | ID: mdl-11279519

ABSTRACT

Variation in the CYP3A enzymes, which act in drug metabolism, influences circulating steroid levels and responses to half of all oxidatively metabolized drugs. CYP3A activity is the sum activity of the family of CYP3A genes, including CYP3A5, which is polymorphically expressed at high levels in a minority of Americans of European descent and Europeans (hereafter collectively referred to as 'Caucasians'). Only people with at least one CYP3A5*1 allele express large amounts of CYP3A5. Our findings show that single-nucleotide polymorphisms (SNPs) in CYP3A5*3 and CYP3A5*6 that cause alternative splicing and protein truncation result in the absence of CYP3A5 from tissues of some people. CYP3A5 was more frequently expressed in livers of African Americans (60%) than in those of Caucasians (33%). Because CYP3A5 represents at least 50% of the total hepatic CYP3A content in people polymorphically expressing CYP3A5, CYP3A5 may be the most important genetic contributor to interindividual and interracial differences in CYP3A-dependent drug clearance and in responses to many medicines.


Subject(s)
Cytochrome P-450 Enzyme System/genetics , Polymorphism, Single Nucleotide , Promoter Regions, Genetic , Alleles , Alternative Splicing , Cytochrome P-450 CYP3A , Humans , Molecular Sequence Data , Racial Groups
6.
Pharm Res ; 17(7): 803-10, 2000 Jul.
Article in English | MEDLINE | ID: mdl-10990198

ABSTRACT

PURPOSE: To develop model polarized cell systems expressing cytochrome P4503A4. NADPH P450 reductase, and P-glycoprotein (Pgp). METHODS: LLC-PK1 and derivative L-MDR1 cells stably expressing Pgp, the product of the multidrug resistance gene (MDR1), were transfected stably using either a mammalian neomycin selectable expression vector (CYP3A4-Neo) or an episomal vector based on Epstein-Barr virus (CYP3A4-Hygro). These CYP3A4 expressing cells were compared with LLC-PK1, L-MDR1, or Caco-2 cells transduced with Adenovirus-3A4 vector (Ad3A4) with or without simultaneous Adenovirus-P450 Reductase (AdRed) transduction. Cells were characterized for expression of CYP3A4 protein and CYP3A4 mediated metabolism towards midazolam and testosterone. Analysis of membrane integrity and drug transport assays were performed to determine whether infection with recombinant Ad3A4 +/- AdRed affected Pgp function. RESULTS: The rank order of optimal CYP3A4 expression and activities in LLC-PKI and L-MDR1 cells from highest to lowest was cells cotransduced with Ad3A4 plus AdRed >> Ad3A4 >>> CYP3A4-Hygro > CYP3A4-Neo. Similarly, coexpression of Ad3A4 plus AdRed led to enhanced CYP3A4 mediated metabolism in Caco-2 cells over cells with Ad3A4 alone. Incubation of transwell cultured cells expressing Ad3A4/AdRed with midazolam led to readily detectable metabolite in the medium. In microsomes from Caco-2 and LLC-PK1 cells, each co-transduced with Ad3A4/AdRed, Vmax values for testosterone 6beta-hydroxylase activity ranged from 414 to 1350 pmoles/min/mg, respectively. For either Caco-2 or LLC-MDR1 cells, TEER values and the rate of apical to basal and basal to apical transport of vinblastine or digoxin were similar in cells with and without Ad3A4/Red transduction. CONCLUSIONS: Polarized cellular systems coexpressing Ad3A4, AdRed, and the MDR1/Pgp transporter were developed and characterized. The results document the utility of these polarized model systems for simultaneous drug transport/drug metabolism studies. Since the experimental approach can be adapted to study the interplay of multiple enzyme/ transporting systems, it may find significant application as a screening tool for the pharmaceutical industry and as a more basic research tool to study the kinetics of intestinal drug bioavailability.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Caco-2 Cells/metabolism , Cytochrome P-450 Enzyme System/metabolism , LLC-PK1 Cells/metabolism , Mixed Function Oxygenases/metabolism , ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , Animals , Cytochrome P-450 CYP3A , Cytochrome P-450 Enzyme System/genetics , Humans , Mixed Function Oxygenases/genetics , NADPH-Ferrihemoprotein Reductase/genetics , NADPH-Ferrihemoprotein Reductase/metabolism , Swine , Transfection/methods
7.
Drug Metab Dispos ; 28(3): 268-78, 2000 Mar.
Article in English | MEDLINE | ID: mdl-10681370

ABSTRACT

Cytochrome P-4503A, CYP2B, and P-450 reductase are induced by glucocorticoids, antiglucocorticoids such as pregnenolone 16alpha-carbonitrile, and drugs such as rifampin and phenobarbital. Although the pregnane X receptor is reported to mediate steroid and drug activation of CYP3A via a conserved cis-element in CYP3A genes, discrepancies exist between the induction of the endogenous CYP3A genes and the activation of the pregnane X receptor. It is a formal possibility that the glucocorticoid receptor may account for some of these discrepancies. To determine the requirement in vivo of the glucocorticoid receptor in expression of CYP3A and CYP2B, we compared the induction of these proteins in the livers of normal mice and mice with a targeted mutation in the glucocorticoid receptor. Mice lacking the glucocorticoid receptor show no difference in constitutive hepatic expression of CYP3A but show a decrease in the level of CYP2B. Glucocorticoid receptor-deficient mice challenged with either dexamethasone or pregnenolone 16alpha-carbonitrile failed to induce CYP2B proteins, whereas CYP2B was readily induced in (+/+) mice. In contrast, CYP3A and P-450 reductase proteins were induced by either inducer in wild-type and glucocorticoid receptor-null mice. Similarly, rifampin induced CYP3A in either wild-type or glucocorticoid receptor-null mice. Despite reports that rifampin is a nonsteroidal ligand for the human glucocorticoid receptor, rifampin failed to induce tyrosine aminotransferase in mice regardless of glucocorticoid receptor genotype, and rifampin did not compete for ligand binding to either mouse or human glucocorticoid receptor. Phenobarbital induced CYP3A, CYP2B, and P-450 reductase in all mice, but the amplitude of induction was diminished 37% in glucocorticoid receptor-null mice. Thus, there are distinctly different essential requirements of CYP3A, CYP2B, and P-450 reductase genes for the glucocorticoid receptor in their induction by steroids and drugs.


Subject(s)
Cytochrome P-450 Enzyme System/metabolism , Liver/drug effects , NADPH-Ferrihemoprotein Reductase/metabolism , Receptors, Glucocorticoid/physiology , Steroids/pharmacology , Aldosterone/metabolism , Animals , Binding, Competitive , Cell Line , Cytochrome P-450 Enzyme System/genetics , Dexamethasone/pharmacology , Enzyme Induction/drug effects , Female , Gene Expression Regulation, Enzymologic/drug effects , Humans , Isoenzymes/genetics , Isoenzymes/metabolism , Liver/enzymology , Male , Mice , Mice, Inbred C57BL , Mice, Inbred Strains , Mice, Mutant Strains , Microsomes, Liver/drug effects , Microsomes, Liver/enzymology , NADPH-Ferrihemoprotein Reductase/genetics , Pregnenolone Carbonitrile/pharmacology , RNA, Messenger/drug effects , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rabbits , Receptors, Glucocorticoid/genetics , Rifampin/metabolism , Rifampin/pharmacology , Tritium
8.
Mol Pharmacol ; 57(1): 188-97, 2000 Jan.
Article in English | MEDLINE | ID: mdl-10617694

ABSTRACT

We hypothesized that the drug efflux protein P-glycoprotein (Pgp), the product of the multidrug resistance gene MDR1, might influence hepatic expression of CYP3A or other cytochromes P-450 (P-450s) because Pgp can transport endogenous regulators of these cytochromes. We began with variants of a CF-1 mouse strain containing a defective mdr1a gene that is inherited in a Mendelian fashion. The amount of CYP3A protein in liver was inversely related to the gene dose of the normal mdr1a allele in these mice. mdr1a knockout mice of either mixed (FVB x 129/Ola) or pure FVB genetic background and housed in Amsterdam display an increased expression of CYP2B and CYP3A proteins. However, because mdr1a ablation causes a compensatory increase in hepatic mdr1b (which can efflux intracellular glucocorticoids), we reasoned that mdr1b might mask the overall effect of mdr1a absence on P-450 gene expression. Targeted inactivation of the mdr1b gene increased P-450 expression, but the effect was modest compared with mdr1a ablation. Mice nullizygous for both mdr1a and mdr1b-type Pgps and kept in Amsterdam had dramatically increased levels of CYP3A protein as well as other P-450s examined and of the electron donor P-450 reductase. Consistent with the protein results, CYP3A catalytic activity measured as midazolam 1'- and 4-hydroxylation in liver microsomes from these knockout mice revealed a rank order of activities with mdr1a/1b > mdr1a > mdr1b > (+/+) mice. In contrast to results in mice housed in Amsterdam, in the genetically identical mdr1a or mdr1a/1b (-/-) male mice housed in the United States, hepatic P-450 expression was unaffected by mdr1 genotype or actually showed a slight decrease in mdr1a (-/-) mice. These results provide a revealing picture of mdr1-type Pgp as an upstream regulator of hepatic P-450 expression, and demonstrate that these pharmacologically relevant phenotypes in knockout mice depend not only on the genetic make-up of the mice but also on the environment.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Aryl Hydrocarbon Hydroxylases , Cytochrome P-450 Enzyme System/biosynthesis , Liver/enzymology , ATP Binding Cassette Transporter, Subfamily B, Member 1/deficiency , ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , Animals , Cytochrome P-450 CYP3A , Cytochrome P-450 Enzyme System/genetics , Female , Gene Expression Regulation, Enzymologic , Male , Mice , Mice, Knockout , NADPH-Ferrihemoprotein Reductase/biosynthesis , NADPH-Ferrihemoprotein Reductase/genetics , Netherlands , Oxidoreductases, N-Demethylating/biosynthesis , Oxidoreductases, N-Demethylating/genetics , United States
9.
Mol Pharmacol ; 54(6): 1113-7, 1998 Dec.
Article in English | MEDLINE | ID: mdl-9855641

ABSTRACT

The pregnenolone X receptor (PXR), a new member of the nuclear hormone receptor superfamily, was recently demonstrated to mediate glucocorticoid agonist and antagonist activation of a hormone response element spaced by three nucleotides (DR-3) within the rat CYP3A23 promoter. Because many other steroids and xenobiotics can up-regulate CYP3A23 expression, we determined whether some of these other regulators used PXR to activate the CYP3A23 DR-3. Transient co-transfection of LLC-PK1 cells with (CYP3A23)2-tk-CAT and mouse PXR demonstrated that the organochlorine pesticides transnonachlor and chlordane and the nonplanar polychlorinated biphenyls (PCBs) each induced the CYP3A23 DR-3 element, and this activation required PXR. Additionally, this study found that PXR is activated to induce (CYP3A23)2-tk-CAT by antihormones of several steroid classes including the antimineralocorticoid spironolactone and the antiandrogen cyproterone acetate. These studies reveal that PXR is involved in the induction of CYP3A23 by pharmacologically and structurally distinct steroids and xenobiotics. Moreover, PXR-mediated PCB activation of the (CYP3A23)2-tk-CAT may serve as a rapid assay for effects of nonplanar PCBs.


Subject(s)
Androgen Antagonists/pharmacology , Aryl Hydrocarbon Hydroxylases , Cyproterone Acetate/pharmacology , Cytochrome P-450 Enzyme System/genetics , Environmental Pollutants/pharmacology , Mineralocorticoid Receptor Antagonists/pharmacology , Receptors, Cytoplasmic and Nuclear/drug effects , Receptors, Steroid/drug effects , Spironolactone/pharmacology , Xenobiotics/pharmacology , Animals , Cell Line , Cell Nucleus/metabolism , Cytochrome P-450 CYP3A , Cytochrome P-450 Enzyme System/chemistry , Enzyme Activation , Mice , Pregnane X Receptor , Pregnenolone/metabolism , Promoter Regions, Genetic , Rats , Receptors, Cytoplasmic and Nuclear/genetics , Receptors, Cytoplasmic and Nuclear/metabolism , Receptors, Steroid/genetics , Receptors, Steroid/metabolism , Response Elements/drug effects , Swine , Transcription, Genetic , Transfection
10.
J Bacteriol ; 180(12): 3209-17, 1998 Jun.
Article in English | MEDLINE | ID: mdl-9620973

ABSTRACT

Pseudomonas aeruginosa a-type strains produce flagellin proteins which vary in molecular weight between strains. To compare the properties of a-type flagellins, the flagellin genes of several Pseudomonas aeruginosa a-type strains, as determined by interaction with specific anti-a monoclonal antibody, were cloned and sequenced. PCR amplification of the a-type flagellin gene fragments from five strains each yielded a 1.02-kb product, indicating that the gene size is not likely to be responsible for the observed molecular weight differences among the a-type strains. The flagellin amino acid sequences of several a-type strains (170,018, 5933, 5939, and PAK) were compared, and that of 170,018 was compared with that of PAO1, a b-type strain. The former comparisons revealed that a-type strains are similar in amino acid sequence, while the latter comparison revealed differences between 170,018 and PAO1. Posttranslational modification was explored for its contribution to the observed differences in molecular weight among the a-type strains. A biotin-hydrazide glycosylation assay was performed on the flagellins of three a-type strains (170,018, 5933, and 5939) and one b-type strain (M2), revealing a positive glycosylation reaction for strains 5933 and 5939 and a negative reaction for 170,018 and M2. Deglycosylation of the flagellin proteins with trifluoromethanesulfonic acid (TFMS) confirmed the glycosylation results. A molecular weight shift was observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis for the TFMS-treated flagellins of 5933 and 5939. These results indicate that the molecular weight discrepancies observed for the a-type flagellins can be attributed, at least in part, to glycosylation of the protein. Anti-a flagellin monoclonal antibody reacted with the TFMS-treated flagellins, suggesting that the glycosyl groups are not a necessary component of the epitope for the human anti-a monoclonal antibody. Comparisons between a-type sequences and a b-type sequence (PAO1) will aid in delineation of the epitope for this monoclonal antibody.


Subject(s)
Flagellin/chemistry , Flagellin/genetics , Genes, Bacterial , Pseudomonas aeruginosa/chemistry , Pseudomonas aeruginosa/genetics , Amino Acid Sequence , Antibodies, Monoclonal , Cloning, Molecular , Epitopes/chemistry , Flagellin/immunology , Glycosylation , Humans , Molecular Sequence Data , Molecular Weight , Polymerase Chain Reaction , Pseudomonas aeruginosa/classification , Sequence Homology, Amino Acid
12.
Haematologia (Budap) ; 22(3): 189-93, 1989.
Article in English | MEDLINE | ID: mdl-2583599

ABSTRACT

A traditional esterolytic assay, a commercial chromogenic assay and an in-house chromogenic assay were compared for suitability in estimating C 1 esterase inhibitor (C 1 INH) activity in plasma and fractions obtained during preparation of a clinical concentrate of C 1 INH. The esterolytic and both of the chromogenic assays showed similar C 1 INH activity in unfractionated plasma samples. However, the esterolytic assay was significantly influenced by the buffer pH and concentration in fractionated plasma samples, and this often resulted in a significant overestimation of C 1 INH in some fractions. Similarly, the commercial C 1 INH assay kit did not always reliably estimate C 1 INH activity in fractions derived from plasma. A modified in-house functional assay was found to be more reliable in measuring C 1 INH in fractionated plasma samples. There was a good correlation between this activity assay and immunological assay estimates of C 1 INH in fractions derived from plasma.


Subject(s)
Complement C1 Inactivator Proteins/analysis , Humans , Immunoassay , Reagent Kits, Diagnostic , Tyrosine
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