Biomedical subjects

L C Chang

Publications and source records attributed to L C Chang.

At least 19 recordsLinked to original sources

PubMed2006

Diffusion tensor imaging in hydrocephalus: initial experience.

PURPOSE AND BACKGROUND: Diffusion tensor imaging (DTI) is an MR imaging-based technique that provides an in vivo tool for visualization of white matter tracts. In this preliminary study, we used this technique to investigate the diffusion characteristics of white matter tracts in patients with hydrocephalus before and after surgery and compared them with age-matched volunteers. MATERIALS AND METHODS: Seven patients with different types of acute hydrocephalus (defined by acute clinical signs of increased intracranial pressure and imaging evidence of enlarged ventricles) underwent MR imaging including a DTI protocol before and after surgery for shunt placement/revision or ventriculostomy. Eight age-matched healthy subjects served as a control group. The DTI was acquired in a clinical setting that included 6 gradient directions with a b value of 1000 s/mm(2). RESULTS: Before surgery, in fiber systems lateral to the ventricles (corona radiata), the diffusion parallel to the fibers was increased (+10%) and the diffusion perpendicular to the fibers was decreased (-25%) in all patients, resulting in an overall increase in the fractional diffusion anisotropy (FA, +28%). Following surgery, the FA values approached those of control values in all except 1 patient. In the corpus callosum, the presurgery FA values in patients with hydrocephalus (HCP) were lower than those of control values, and no significant changes were seen following surgery. CONCLUSIONS: DTI can distinguish the compression characteristics of white matter before and after surgery in patients with HCP. At the acute stage of the disease, DTI characteristics point to white matter compression as a possible cause of the observed changes.

Acute Disease↗

PubMed2004

Splenic tumour: a clinicopathological study.

Splenic tumours are occasionally found during routine physical check-ups or elective abdominal image studies. Histologically, most splenic tumours are of benign vascular origin. To avoid unnecessary surgery for asymptomatic patients with benign splenic tumours and clarify the clinicopathological features of spleen tumours, this study gathered 44 cases of primary or isolated metastatic spleen tumours confirmed by pathology from surgery specimens or biopsies. The differences in clinicopathological features and image presentations between benign and malignant spleen tumour were investigated. Thirty-two cases involved benign tumours while 12 cases were malignant. Among the benign tumours, vascular originating tumours were most common (with 14 cases of cavernous haemangiomas, 13 cases of lymphangioma, three cases of lymphangiohaemangioma and one case of Littoral cell angioma). Notably, one, case of inflammatory pseudotumour because of Schistosoma parasite infection was also noted. Among the malignant tumours, there were four cases of angiosarcomas with vascular endothelium origins, as well as lymphomas and six metastatic tumours. Image studies were non-specific. Image study alone is an inadequate basis for making differential diagnoses between benign and malignant tumours. Instead, pathological studies are required for a final diagnosis. Using previous studies and this investigation, fine needle aspiration biopsy of spleen tumours with the help of ultrasonic or computed tomography appears a safe and effective method for obtaining biopsy specimens. Splenectomy is recommended only for patients with malignancies or complications such as intractable abdominal pain, coagulopathy or tumour rupture with an unstable haemodynamic state.

Adolescent↗

PubMed2001

Inhibition of extracellular Ca(2+) entry by YC-1, an activator of soluble guanylyl cyclase, through a cyclic GMP-independent pathway in rat neutrophils.

The effects of a soluble guanylyl cyclase (sGC) activator, 3-(5'-hydroxymethyl-2'-furyl)-1-benzyl indazole (YC-1), on formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated [Ca(2+)](i) elevation in rat neutrophils were examined. YC-1 produced a concentration-dependent inhibition of [Ca(2+)](i) elevation. Pretreatment of neutrophils with YC-1 did not enhance its inhibitory effect. YC-1 also inhibited the [Ca(2+)](i) changes caused by ionomycin. In a biphasic model, measuring the [Ca(2+)](i) stimulation by fMLP in a Ca(2+)-free medium followed by reintroduction of Ca(2+), YC-1 mainly affected Ca(2+) influx. YC-1 also inhibited active and passive Mn(2+) influx, and this inhibitory effect was not attenuated by the sGC inhibitor 6-anilino-5,8-quinolinequinone (LY83583). Sodium nitroprusside did not affect the fMLP-stimulated [Ca(2+)](i) changes. Pretreatment of neutrophils with the cyclic GMP-dependent protein kinase inhibitor 8-(4-chlorophenylthio) guanosine-3',5'-monophosphorothioate, Rp-isomer (Rp-8-pCPT-cGMPS), LY83583, the protein phosphatase 2B inhibitor cyclosporin A, or the protein kinase inhibitor staurosporine did not attenuate the inhibition of [Ca(2+)](i) by YC-1. YC-1 inhibited the fMLP-stimulated protein tyrosine phosphorylation. These results indicate that cyclic GMP does not play an important role in the regulation of [Ca(2+)](i) in rat neutrophils. Inhibition of fMLP-stimulated [Ca(2+)](i) changes by YC-1 is mainly via the blockade of Ca(2+) entry through the inhibition of tyrosine kinase activity, but not the stimulation of protein kinase C and protein phosphatase 2B.

Animals↗

PubMed2001

The reappraisal of nephrocalcin--its role in the inhibition of calcium oxalate crystal growth and interaction with divalent metal ions.

Nephrocalcin (NC) is a potent crystal growth inhibitor of calcium oxalate monohydrate. However, the sequence is undefined owing to its multiple glycosylations. Although there have been many biochemical studies of the binding capacity of calcium, the study of the function of the domain is still deferred. By using S-200 gel filtration and Mono Q ion-exchange chromatographic procedures, NC can be purified without further treatment for the removal of urobilirubin. The kinetic study of crystal growth in calcium oxalate monohydrate is performed using a seed slurry system. NC was cut into two peptides through interaction with copper ion and ascorbic acid. The interaction site of the copper ion is presumed to be located between 8 and 6 kDa of molecular weight in NC. The data suggest that divalent metal ions may be involved in the calcium oxalate crystallization through interaction with NC. The role of ascorbic acid in the formation of urinary stones should be reappraised for its association in the redox reaction, with resultant protein digestion in the presence of copper ions.

Adult↗

PubMed2001

Stability of 4-DMAP in solution.

A stability-indicating reversed-phase performance liquid chromatographic method was developed for the detection of 4-(N,N-dimethylamino)phenol (4-DMAP) and its degradation products under accelerated degradation conditions. The degradation kinetics of 4-DMAP in aqueous solution over a pH range of 1.12-6.05 and its stability in solutions based on propylene glycol or polyethylene glycol 400 were investigated. The observed rate constants were shown to follow, apparent first-order kinetics in all cases. The pH rate profile shows that maximum stability of 4-DMAP was observed in the pH range 2.0 to 3.0. Acid/base catalysis of 4-DMAP was not affected by systems of various ionic strengths. Incorporation of nonaqueous propylene glycol or polyethylene glycol 400 in the pH 3.05 solution of 4-DMAP showed an increase in the stability at 55 degrees C +/- 0.5 degrees C.

4-Aminopyridine↗

PubMed2001

Low molecular weight protamine (LMWP) as nontoxic heparin/low molecular weight heparin antidote (I): preparation and characterization.

Low molecular weight protamine (LMWP) appears to be a promising solution for heparin neutralization without the protamine-associated catastrophic toxic effects. The feasibility of this hypothesis was proven previously by using a peptide mixture produced from proteolytic digestion of protamine. To further examine the utility of this compound as an ultimate nontoxic protamine substitute, detailed studies on the purification and characterization of LMWP including the precise amino acid sequence, structure-function relationship, and possible mechanism were conducted. A number of LWMP fragments, composed of highly cationic peptides with molecular weights ranging from 700 to 1900 d, were prepared by digestion of native protamine with the protease thermolysin. These fragments were fractionated using a heparin affinity chromatography, and their relative binding strengths toward heparin were elucidated. Five distinct fractions were eluted at NaCl concentration ranging from 0.4 to 1.0 M and were denoted as TDSP1 to TDSP5, in increasing order of eluting ionic strength. Among these 5 fractions, TDSP4 and TDSP5 contained 3 LMWP peptide fragments, and they were found to retain the complete heparin-neutralizing function of protamine. By using a peptide mass spectrometry (MS) fingerprint mapping technique, the amino acid sequences of the microheterogeneous LMWP fragments in all these 5 elution fractions were readily identified. A typical structural scaffold made by arginine clusters in the middle and nonarginine residues at the N-terminal of the peptide sequence was observed for all these LMWP fragments. By aligning the sequences with the potency in heparin neutralization of these LMWP fragments, it was found that retention of potency similar to that of protamine required the presence of at least 2 arginine clusters in the LMWP fragments; such as the sequence of VSRRRRRRGGRRRR seen in the most potent LMWP fraction-TDSP5. The above finding was further validated by using a synthetic LMWP analogue-CRRRRRRR-and it was found that its heparin-neutralizing ability was increased by changing from a monomeric to a dimeric structure of this analogue peptide. Based on these results, the structural requirement for a compound to function as an effective heparin antidote and the possible mechanism involved in heparin neutralization were established.

Anticoagulants↗

PubMed2001

Low molecular weight protamine (LMWP) as nontoxic heparin/low molecular weight heparin antidote (II): in vitro evaluation of efficacy and toxicity.

Patients undergoing anticoagulation with heparin or low molecular weight heparin (LMWH) require a superior antidote that possesses more selective biological actions and a better safety profile than protamine. We had previously developed 2 low molecular weight protamine (LMWP) fractions (TDSP4 and TDSP5) from thermolysin-digested protamine as potential nontoxic, heparin-neutralizing agents. In this, the second article in this series, studies focused on in vitro evaluation of heparin/LMWH-neutralizing efficacy and putative toxicity. These LMWP fractions, particularly TDSP5, were effective and fully capable of neutralizing a broad spectrum of heparin-induced anticoagulant activities (ie, aPTT, anti-Xa, and anti-IIa activities). Additionally, these LMWP fractions could neutralize the activities of commercial LMWH. As assessed by the anti-Xa assay, TDSP5 was as effective as, although less potent than, protamine in reversing the activity of Mono-Embolex (molecular weight 5000-7000) and 2 other different sizes (molecular weight of 3000 and 5000 d) of LMWH preparations. Furthermore, compared with protamine, TDSP5 exhibited a much-reduced toxicity and thus an improved safety profile, as reflected by its reduced ability to activate the complement system and cross-react with the antiprotamine antibodies, which are 2 primary indices of protamine toxicity.

Animals↗

PubMed2001

Low molecular weight protamine as nontoxic heparin/low molecular weight heparin antidote (III): preliminary in vivo evaluation of efficacy and toxicity using a canine model.

Heparin employed in cardiovascular surgeries often leads to a high incidence of bleeding complications. Protamine employed in heparin reversal, however, can cause severe adverse reactions. In an attempt to address this clinical problem, we developed low molecular weight protamine (LMWP) as a potentially effective and less toxic heparin antagonist. A homogeneous 1880-d peptide fragment, termed LMWP-TDSP5 and containing the amino acid sequence of VSRRRRRRGGRRRR, was derived directly from protamine by enzymatic digestion of protamine with thermolysin. In vitro studies demonstrated that TDSP5 was capable of neutralizing various anticoagulant functions of both heparin and commercial low molecular weight heparin preparations. In addition, TDSP5 exhibited significantly reduced crossreactivity toward mouse sera containing antiprotamine antibodies. TDSP5 showed a decrease in its potential in activating the complement system. All of these findings suggested the possibility of markedly reduced protamine toxicity for TDSP5. In this article, we conducted preliminary in vivo studies to further demonstrate the feasibility and utility of using LMWP as a nontoxic clinical protamine substitute. Dogs were chosen as test animals because they were known to magnify the typical human response to protamine. By using a full spectra of biological and clinical assays for heparin, including the anti-IIa and anti-Xa chromogenic assays and the activated partial, thromboplastin time and TCT clotting assays, TDSP5 showed that it could completely neutralize all these different anticoagulant functions of heparin in dogs. Although administration of protamine in dogs produced a significant reduction in mean arterial blood pressure (-14.9 mm Hg) and elevation in pulmonary artery systolic pressure (+5.0 mm Hg), the use of TDSP5 in dogs did not elicit any statistically significant change in any of the variables measured. Furthermore, the use of LMWP also significantly reduced the protamine-induced transient thrombocytopenic and granulocytopenic responses. The white blood cell counts and platelet counts decreased to 82.1% and 60.0% of baseline, respectively, in dogs given intravenous protamine compared to 97.8% and 88.6% of baseline in dogs receiving TDSP5. These preliminary findings indicated that LMWP could potentially provide an effective and safe means to control both heparin- and protamine-induced complications.

Animals↗

PubMed2001

Signal transduction pathways for activation of extracellular signal-regulated kinase by arachidonic acid in rat neutrophils.

Phosphorylation of extracellular signal-regulated kinase (ERK) in response to arachidonic acid (AA) was rapid and transient, peaking at 1 min and disappearing after 3 min, and it was accompanied by an increase in ERK activity in rat neutrophils. We examined the upstream regulation of AA-stimulated ERK activation using one of the following signaling pathway inhibitors to pretreat rat cells: the ERK kinase inhibitor U0126 or PD98059, the G(i/o) inhibitor pertussis toxin (PTX), the tyrosine kinase inhibitor genistein, the phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin or LY294002, the Ca2+ chelator 1,2-bis(O-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid, or the phospholipase C (PLC) inhibitor U73122. All of these inhibitors attenuated AA-induced ERK activation. Activation of ERK was also effectively attenuated by the cyclooxygenase and lipoxygenase inhibitor BW755C and by the leukotriene biosynthesis inhibitor MK886, but the cyclooxygenase inhibitor indomethacin did not attenuate ERK activation. After exposing cells to three distinct protein kinase C (PKC) inhibitors, we found that Gö6976 significantly attenuated ERK phosphorylation but potentiated ERK activity. Neither Gö6983 nor GF109203X affected AA-induced responses. These data suggest that the lipoxygenase metabolite(s) produced mediates AA-stimulated ERK activation and that this effect is upstream regulated by PT-sensitive G protein, non-receptor tyrosine kinase, PI3K, and PLC/Ca2+ signaling pathways in rat neutrophils.

4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyraz↗

PubMed2000

Anticancer activity evaluation of the solanum glycoalkaloid solamargine. Triggering apoptosis in human hepatoma cells.

Solamargine, an herbal and molluscicidal medicine derived from Solanum incanum, is a steroidal alkaloid glycoside. To characterize the anticancer mechanism of solamargine on human hepatoma cells (Hep3B), changes of cell morphology, DNA content, and gene expression of cells after solamargine treatment were studied. The appearance in solamargine-treated cells of chromatin condensation, DNA fragmentation, and a sub-G(1) peak in a DNA histogram suggests that solamargine induces cell death by apoptosis. The maximum number of dead Hep3B cells was detected within 2 hr of incubation with constant concentrations of solamargine, and no further cell death was observed after an extended incubation with solamargine, indicating that the action of solamargine was irreversible. To determine the susceptibility of cell phases to solamargine-mediated apoptosis, Hep3B cells were synchronized at defined cell cycles by cyclosporin A, colchicine, and genistein, followed by solamargine treatment. The IC(50) values of solamargine for control, G(0)/G(1)-, M-, and G(2)/M-synchronized Hep3B cells were 5.0, > 10, 3.7, and 3.1 microg/mL, implying that cells in the G(2)/M phases are relatively susceptible to solamargine-mediated apoptosis. In addition, a parallel up-regulation of tumor necrosis factor receptor (TNFR)-I and -II on Hep3B cells was detected after solamargine treatment, and the solamargine-mediated cytotoxicity could be neutralized with either TNFR-I or -II specific antibody. Therefore, these results reveal that the actions of TNFR-I and -II on Hep3B cells may be independent, and both are involved in the mechanism of solamargine-mediated apoptosis.

Antibodies↗

PubMed2000

Activity-guided isolation of constituents of Cerbera manghas with antiproliferative and antiestrogenic activities.

Two new cardenolides, (-)-14-hydroxy-3beta-(3-O-methyl-6-deoxy-alpha-L-rhamnosyl)-11a lpha, 12alpha-epoxy-(5beta,14beta,17betaH)-card-20 (22)-enolide (1), (-)-14-hydroxy-3beta-(3-O-methyl-6-deoxy-alpha-L-glucopyranosyl)-11al pha,12alpha-epoxy-(5beta,14beta,17betaH)-card -20(22)-enolide (2), and a known cardenolide, (-)-17beta-neriifolin (3), were isolated from the roots of Cerbera manghas as antiproliferative and antiestrogenic principles when evaluated against a human colon cancer cell line (Col2) and the Ishikawa cell line, respectively. Two known lignans, (-)-olivil (4) and (-)-cycloolivil (5), were also isolated but were inactive in the assay systems used.

Antineoplastic Agents, Phytogenic↗

PubMed2000

Quantitative determination of the expression of xeroderma pigmentosum F gene in human nonmelanoma skin cancers.

Nonmelanoma skin cancers (NMSC) has been evidenced with an impaired function in nucleotide excision repair (NER). However, malfunction of NER elements in NMSC has not been identified. Xeroderma pigmentosum F (XPF) is an essential subunit in NER and functions as a 5'-incision enzyme when repairing damaged DNA. So far, neither XPF's protein nor antibody is commercially available. To explore the expression of XPF in NMSC, the gene was determined by quantitative reverse transcription-polymerase chain reaction (RT-PCR). All the designed primers specifically amplified XPF cDNA as demonstrated by nested PCR, and one set of the primers was mimic constructed to form a controlled cDNA for the semiquantification of XPF gene in NMSC. The results indicated that the quantities of XPF expression of BCC and SCC specimens were approximately 57.0 and 76.4% less than that of normal skins, respectively. This paper indicates that the decrease expression of XPF gene may be one of mechanisms for impaired NER in NMSC, and the feasible and quantitative primers used in the experiments may explore the study of XPF in etiology of carcinogenesis.

Base Sequence↗

PubMed2000

Activation of p38 mitogen-activated protein kinase by formyl-methionyl-leucyl-phenylalanine in rat neutrophils.

The signaling pathways leading to p38 mitogen-activated protein kinase (MAPK) activation in formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated rat neutrophils were examined. Immunoblot analysis with antibodies against a phosphorylated form of p38 MAPK showed that fMLP-stimulated p38 MAPK activation was dependent on a pertussis toxin-sensitive G protein. Two phosphatidylinositol 3-kinase inhibitors, wortmannin and 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), did not affect the p38 MAPK activation. Phosphorylation of p38 MAPK was concentration dependently attenuated by a tyrosine kinase inhibitor, genistein, and by a Ca(2+)-dependent protein kinase C inhibitor, 13-cyanoethyl-12-methyl-6,7,12,13-tetrahydroindolo[2,3-a]pyrrolo[3 , 4-c]carbazole-7-one (Gö6976). However, the protein kinase C inhibitors with a broader spectrum, 2-[1-(3-dimethylaminopropyl)-5-methoxy-1H-indol-3-yl]-3-(1H-indol-3-y l)-maleimide (Gö6983) and 2-[1-(3-dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3-yl)-maleimi de (GF109203X), had no inhibitory effect. fMLP-stimulated p38 MAPK phosphorylation was also reduced in cells pretreated with a phospholipase C inhibitor, 1-[6-((17beta-3-methoxyestra-1,3, 5(10)-trien-17-yl)amino)hexyl]-1H-pyrrole-2,5-dione (U73122), or preloaded with an intracellular Ca(2+) chelator, 1, 2-bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA). We conclude that phosphorylation of p38 MAPK by fMLP stimulation in rat neutrophils is dependent on G(i/o) protein, nonreceptor tyrosine kinase, phospholipase C/Ca(2+), and probably Ca(2+)-dependent protein kinase C pathways.

Animals↗

PubMed2000

Absolute configuration of novel bioactive flavonoids from Tephrosia purpurea.

[structure: see text] Three novel flavonoids, (+)-tephrorins A (1) and B (2) and (+)-tephrosone (3), were isolated from Tephrosia purpurea. Their structures were elucidated by NMR spectral analysis, and their absolute configurations were determined by Mosher ester methodology. Compounds 1 and 2 are flavanones containing an unusual tetrahydrofuran moiety. Compounds 1-3 were evaluated for their potential cancer chemopreventive properties using a cell-based quinone reductase induction assay.

Animals↗

PubMed2000

Aromatase inhibitors from Isodon excisus var. coreanus.

The diethyl ether extract of Isodon excisus var. coreanus exhibited significant inhibitory activity in aromatase assay. Bioactivity-guided fractionation of the extract led to the isolation of three active compounds: inflexin (ent-1alpha-hydroxy-3beta,6a-diacetoxykaur-16-en-11,15-dione ) (1), ursolic acid (2), and ursolic acid 3-O-acetate (3).

Anticarcinogenic Agents↗

PubMed2000

Bioactive constituents of Thuja occidentalis.

An ethyl acetate-soluble extract of the combined leaves and twigs of Thuja occidentalis was found to inhibit 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced ornithine decarboxylase (ODC) in cultured mouse epidermal ME 308 cells. Bioassay-guided fractionation of this extract led to the isolation of six active constituents (1-6), namely, (+)-7-oxo-13-epi-pimara-14,15-dien-18-oic acid (1), (+)-7-oxo-13-epi-pimara-8,15-dien-18-oic acid (2), (+)-isopimaric acid (3), (1S,2S,3R)-(+)-isopicrodeoxypodophyllotoxin (4), (-)-deoxypodophyllotoxin (5), and (-)-deoxypodorhizone (6). Compounds 1 and 4 are new natural products, and their structures and stereochemistry were determined using spectroscopic methods. Compounds 1-6 were evaluated for inhibition of the transformation of murine epidermal JB6 cells, inhibition of ornithine decarboxylase induction with murine epidermal ME 308 cells, and cytotoxic activity against KB cells.

Animals↗

PubMed2000

The upstream regulation of p38 mitogen-activated protein kinase phosphorylation by arachidonic acid in rat neutrophils.

The signal transduction pathways activated by arachidonic acid that lead to p38 mitogen-activated protein kinase (MAPK) activation in neutrophils remains unclear. In this study, selective inhibitors of several signalling pathways were utilized to investigate the mechanisms of activation of p38 MAPK by arachidonic acid in rat neutrophils. Stimulation of p38 MAPK phosphorylation by arachidonic acid and its trifluoromethyl ketone analogue AACOCF3 was transient, peaking at 1 min, and was concentration-dependent. Arachidonic acid-stimulated p38 MAPK phosphorylation was attenuated in cells pretreated with the Gi/o inhibitor (pertussis toxin), but not with the dual cyclooxygenase/lipoxygenase inhibitor (BW755C) or the leukotriene biosynthesis inhibitor (MK886). Tyrosine kinase inhibitor (genistein), but not the extracellular signal-regulated kinase kinase inhibitors (PD98059 and U0126), attenuated the phosphorylation of p38 MAPK by arachidonic acid. Phosphoinositide 3-kinase inhibitors (wortmannin and LY294002) did not affect the arachidonic acid-induced response. After pretreatment of the cells with protein kinase C inhibitors (Gö6976, Gö6983 and GF109203X), only Gö6976 significantly attenuated the phosphorylation of p38 MAPK by arachidonic acid. In addition, phosphorylation of p38 MAPK by arachidonic acid was greatly attenuated by the phospholipase C inhibitor (U73122) and the Ca2+ chelator BAPTA ((1,2-bis-o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid), but not altered by the nitric oxide synthase inhibitor, N-nitro-L-arginine methyl ester. Arachidonic acid did not cause an increase in cellular cyclic GMP level. This study revealed the involvement of pertussis toxin-sensitive G protein, non-receptor tyrosine kinase, phospholipase C/Ca2+, and probably Ca2+-dependent protein kinase C in arachidonic acid-stimulated p38 MAPK activation.

Animals↗

PubMed1999

Electrochemical assay of plasminogen activators in plasma using polyion-sensitive membrane electrode detection.

Two relatively simple electrochemical assay methods suitable for the measurement of plasminogen activators (including urokinase (u-PA), streptokinase (SK), and tissue plasminogen activator (t-PA)) in plasma samples are described. In one approach, the initial rate of decrease in the potentiometric response of a polycation-sensitive membrane electrode toward protamine is monitored after addition of a preincubated reaction mixture containing the sample and exogenous plasminogen. The plasmin formed from plasminogen by the activators catalyzes the decomposition of the arginine-rich protamine substrate, yielding smaller polycationic fragments that are not sensed by the electrode. Alternately, the sample, plasminogen, and protamine can be incubated together, and the remaining protamine in this reaction mixture can be measured at a fixed point in time by placing the electrode into the mixture and recording the electromotive force response. Working curves found with both methods for plasma samples spiked with varying levels of the activators cover the expected therapeutic activity ranges found in the plasma of patients treated with these "clot-busting" drugs.

Blood Chemical Analysis↗