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Biomedical subjects

A Salvi

Publications and source records attributed to A Salvi.

At least 19 recordsLinked to original sources

RNA interference against urokinase in hepatocellular carcinoma xenografts in nude mice.

The serine protease urokinase-type plasminogen activator (u-PA) is overexpressed in hepatocellular carcinoma (HCC) and its expression level is inversely correlated with the patients' survival. The purpose of this study was to examine the effects of vector-based RNA interference (RNAi) of u-PA on the growth of human HCC xenografts in nude mice in order to investigate the role of u-PA in human HCC. Our results showed that the subcutaneous injection of small interfering RNAs (siRNA) u-PA SKHep1C3 stable transfected cells (pS siRNA u-PA) led to a growth delay in xenograft development, compared to those generated from empty vector; the molecular characterization of nodules (carried out by PCR, RT-PCR and immunohistochemical analysis) revealed the presence of plasmid DNA, the u-PA gene expression knockdown, at both mRNA and protein levels, giving evidence of a long-term and target-specific inhibition by vector-based RNAi 11 weeks after cell inoculation. We further studied the effects of u-PA down modulation on extracellular matrix (ECM) proteins evaluating the expression and organization of fibronectin (FN; one of the main ECM proteins). Immunohistochemical and immunofluorescence analysis of FN revealed FN fibrils in pS siRNA u-PA xenografts and in pS siRNA u-PA cells, thus identifying the FN fibril organization as a downstream effect of u-PA knockdown in this system.

Animals↗

Long term effectiveness of intravenous immunoglobulin in Churg-Strauss syndrome.

OBJECTIVE: To study the long term effectiveness of intravenous immunoglobulin and plasmapheresis associated with prednisone and cyclophosphamide in Churg-Strauss syndrome. SUBJECTS: and methods: We studied 18 subjects with new onset Churg-Strauss syndrome. All received the "standard" treatment based on prednisone (1 mg/kg/day for 1 month and then slowly tapered) and cyclophosphamide (2 mg/kg/day for 6 months in severe cases). In nine patients, synchronised cycles with plasmapheresis and intravenous immunoglobulin (2 g/kg) were repeated monthly for 6 months and every other month for a further three cycles. Clinical (disease activity monitored by Birmingham vasculitis activity score (BVAS) and damage index (modified Rankin score)) and functional (C reactive protein, blood eosinophil count, and electromyogram-electoneurogram) parameters were collected during treatment and the 3 year follow up period. RESULTS: After 12 months, all patients in the treatment group and four (44%) in the control group were in remission. At the end of the 3 year follow up period, we documented significant differences in BVAS (p<0.01), global damage (p<0.02), modified Rankin score (p<0.04), and the daily maintenance prednisone dose (p<0.002) between the two groups. We found a tendency towards lower frequency of relapse and incidence of osteoporosis in the treatment group. CONCLUSION: Complete clinical and functional recovery with a long term stable remission and a low incidence of side effects can be achieved by intravenous immunoglobulin associated with plasmapheresis in patients with Churg-Strauss syndrome.

Adult↗

Cyclosporin A and intravenous immunoglobulin treatment in polymyositis/dermatomyositis.

OBJECTIVE: To describe the treatment of polymyositis (PM) and dermatomyositis (DM) with prednisone (PRED) and cyclosporin A (CSA) alone or associated with intravenous immunoglobulin (IVIg) and plasmapheresis (PEX). METHODS: Between 1992 and 1999 CSA and PRED were used to treat 20 patients with idiopathic myositis (12 with DM, eight with PM), diagnosed according to the Bohan and Peter criteria. In patients with refractory or relapsed disease, IVIg was added alone (seven cases) or synchronised with PEX (six cases). A standardised protocol was used to evaluate the patients, and assess disease activity and treatment response. RESULTS: Despite a transient response to PRED and CSA in 16/20 cases, this combination did not induce full remission in 13/20 cases, which led to the IVIg trial with or without PEX. Patients receiving PRED and CSA plus IVIg had a significantly higher probability of maintaining complete remission at the end of the four year follow up period than those treated with PRED and CSA alone (p<0.001). No further benefit was added by the PEX. The presence of arthritis significantly correlated with a poorer response to treatment (p<0.05). Adverse effects were gingival hyperplasia (one patient) and transient renal dysfunction (one). CONCLUSIONS: This open study suggests that combined treatment with PRED, CSA, and IVIg is useful in patients with myositis, even those with refractory or relapsed disease; no increase in the number or type of side effects is seen.

Adult↗

Structural damage to proteins caused by free radicals: asessment, protection by antioxidants, and influence of protein binding.

Oxidative damage to proteins results in biological dysfunctions such as perturbed activity in enzymes, transport proteins, and receptors. Here, we investigated structural damage to proteins induced by free radicals. Structural alterations to lysozyme, human serum albumin (HSA) and beta-lactoglobulin A were monitored by capillary zone electrophoresis. Four well-known antioxidants (quercetin, melatonin, Trolox, and chlorogenic acid) were examined for their ability to inhibit protein damage and to bind to these proteins. Melatonin and chlorogenic acid, which did not bind to any of the three proteins under study, showed scavenging and protective activities well correlated with the amount of free radicals generated. Trolox, which bound only to HSA, was a better protector of HSA than of the two other proteins, indicating that its antioxidant capacity is increased by a shielding effect. Finally, quercetin was a good antioxidant in protecting lysozyme and beta-lactoglobulin A, but its binding to HSA resulted in a pro-oxidant effect that accelerated HSA fragmentation. These results demonstrate that binding of an antioxidant to a protein may potentiate protection or damage depending on the properties of the antioxidant.

Amidines↗

Serum imbalance of cytokines in melanoma patients.

The cytokines interleukin (IL)6 and IL10 appear to be involved in the progression of melanoma because they are secreted by malignant cells and their serum levels are associated with poor survival and with advanced stages of the disease. Antitumour immunity is considered to be a T-cell response, mediated mainly by type 1 cytokines such as IL12 and interferon-gamma (IFNgamma). We evaluated the serum levels of cytokines involved in the host response against tumour (IL12, IFNgamma) and/or the progression of melanoma (IL6, IL10) in 45 melanoma patients with localized and metastatic disease and in 45 controls, using commercially available enzyme-linked immunosorbent assay (ELISA) kits. In the controls, IL6 and IL12 were nearly undetectable, whereas the IL10 and IFNgamma ranges were 0.5-9 pg/ml and 2-4.8 pg/ml, respectively. In the melanoma patients, pathologically high values were found in 44.4% for IL6, in 24.4% for IL10, and in 60% for IL12. Significantly higher values were found for IL6 and IL12, and lower values for IFNgamma. This study highlights a significant difference in serum cytokine profiles between controls and melanoma patients, which is mainly due to the high levels of IL6 and IL12 and the low levels of IFNgamma.

Cytokines↗

A stilbene and dihydrochalcones with radical scavenging activities from Loiseleuria procumbens.

A new dihydrochalcone, 6''-acetylphloridzosid, was isolated from the whole plant of Loiseleuria procumbens (L.) Desv. and identified as 2'-O-(6''-O-acetylglucopyranosyl)-4,4',6'-trihydroxydihydrochal cone by spectroscopic methods. In addition, one stilbene and three other dihydrochalcones were identified as (E)-piceid, phloretin (2',4,4',6'-tetrahydroxydihydrochalcone), phloridzosid (2'-O-glucopyranosyl-4,4',6'-trihydroxydihydrochalcone) and asebotin (2'-O-glucopyranosyl-4'-methoxy-4,6'-dihydroxydihydrochalcone), respectively. Some of these compounds showed scavenging properties towards the 2,2-diphenyl-1-picrylhydrazyl radical and antioxidant properties in a test with lysozyme.

Chalcone↗

A role for mucosal immunity in resistance to HIV infection.

In a recent, thought-provoking novel by Elizabeth McCracken (The Giant's House. Avon Books, New York, 1997), two characters discuss love and its impossibilities. One brashly claims to be "immune to love", explaining the concept to his perplexed interlocutor, "...people become immune to love like they become immune to any disease. Either they had it bad early in life, like chicken pox and that's that; or they keep getting exposed to it in little doses and build up an immunity; or somehow they just don't catch it, something in'em is born resistant. I'm the last type. I'm immune to love and poison ivy". (p. 275) (E. McCracken, The Giant's House. Avon Books, New York, 1997). Substitute the words 'HIV infection' for 'love' and this intriguing metaphor summarizes the state of the art working hypotheses for the phenomenon of resistance to HIV infection in HIV-exposed individuals who, against all odds, do not seroconvert. These hypotheses will be discussed hereafter and particular emphasis will be placed upon a possible role for mucosal immunity in this phenomenon.

Chemokines↗

Monocytes of patients wiht systemic sclerosis (scleroderma spontaneously release in vitro increased amounts of superoxide anion.

It has been suggested that toxic oxygen free radicals can be involved in the pathogenesis of systemic sclerosis (scleroderma) (SSc). Because the cells that contribute to the generation of free radicals are not known, our aim was (i) to evaluate the ability of unmanipulated and phorbol 12-myristate 13-acetate-stimulated monocytes and polymorphonucleate neutrophils of SSc patients to generate superoxide anion (O2*-); and (ii) to investigate whether the O2*- produced by these cells involved the activation of nicotinamide-adenine dinucleotide diphosphate oxidase biochemical pathway. Employing the superoxide dismutase-inhibitable reduction of cytochrome c to evaluate the generation of O2*-, unmanipulated monocytes of SSc patients generated more O2*- than primary Raynaud's phenomenon patients and normal control monocytes (p = 0.0001), and the release was higher in patients with diffuse cutaneous involvement and 5 y or less disease duration (p = 0.02). The involvement of nicotinamide-adenine dinucleotide diphosphate oxidase in the enhanced 02*- production was demonstrated by the finding that the cytosolic components of the enzyme, p47phox and p67phox, were both translocated to the plasma membrane of enriched but otherwise unmanipulated monocytes of SSc patients. The involvement of mitochondrial oxidases was excluded by the lack of inhibition of O2*- production when monocytes were incubated in the presence of rotenone, a mitochondrial oxidase inhibitor. Upon stimulation with phorbol 12-myristate 13-acetate, monocytes of SSc patients produced more O2*- than controls. In SSc patients untreated polymorphonucleate neutrophils generated significantly less O2*- than monocytes (p = 0.0001) and only slightly more than polymorphonucleate neutrophils of primary Raynaud's phenomenon patients and normal controls (p = 0.03). In conclusion, we demonstrate that in patients with scleroderma, unmanipulated and phorbol 12-myristate 13-acetate-stimulated monocytes release in vitro increased amounts of superoxide anion through the activation of nicotinamide-adenine dinucleotide diphosphate oxidase and, thus, contribute to the oxidative stress found in this disease.

Adult↗

Human immunodeficiency virus (HIV)-specific IgA and HIV neutralizing activity in the serum of exposed seronegative partners of HIV-seropositive persons.

The presence and activity of human immunodeficiency virus (HIV)-specific antibodies were analyzed in the sera of 15 sexually exposed seronegative persons who had systemic HIV-specific cell-mediated immunity and IgA-mediated mucosal immunity and in their HIV-infected partners. The HIV-positive subjects had HIV-specific serum IgG and IgA; the seronegative persons had HIV-specific serum IgA in the absence of IgG. Testing of the seronegative persons 1 year after the interruption of at-risk sex showed that no IgG seroconversion had occurred and that HIV-specific IgA serum concentrations had declined. Serum from the HIV-exposed seronegative persons was analyzed for the ability to neutralize primary HIV-1 isolates. Neutralizing activity was detected in 5 of 15 sera and in 2 cases was retained by serum-purified IgA. Thus, the immunologic picture for resistance to HIV infection should include HIV-specific cell-mediated immunity as well as HIV-specific IgA-mediated mucosal and systemic immunity.

Female↗

Undifferentiated connective tissue disease: natural history and evolution into definite CTD assessed in 84 patients initially diagnosed as early UCTD.

Connective tissue diseases (CTDs) are chronic multisystemic inflammatory disorders whose indicative signs or symptoms have a high sensitivity but poor specificity in predicting the evolution into a given CTD. We have analysed 84 consecutive patients initially diagnosed as having an early undifferentiated CTD (early UCTD) with the aim of verifying the evolution into one definite CTD and of evaluating the predictive value of clinical and laboratory parameters. During a 5-year study period, 33 patients developed signs of a full-blown CTD; the highest probability of evolution was in the first 48 months after the onset. Multivariate analysis allowed us to select those variables correlating with evolution into a particular CTD, such as sclerodactyly and oesophageal dysfunction for systemic sclerosis, xerostomia and anti-nuclear antibodies (SS-A pattern) for Sjögren's syndrome, and fever and anti-DNA antibodies for systemic lupus erythematosus. Furthermore, we assessed the prevalence of various clinical and laboratory manifestations, complications and prognosis of those patients diagnosed after a 5-year disease duration period as having a UCTD. In our series, major organs such as the kidney or heart seem to be spared, whereas we detected a relatively high prevalence of endocrine disease of autoimmune origin.

Adolescent↗

Chlamydia pneumoniae antibody response in patients with acute myocardial infarction and their follow-up.

STUDY POPULATIONS: This study concerned the possible relations between seroreactivity to Chlamydia pneumoniae and myocardial infarction. A group of 29 patients with acute myocardial infarction (AMI), 74 members of a healthy control group, and a subgroup of 24 members of a healthy control group matched for age, sex, and coronary risk factors (HCM) were included in the study. In addition, we evaluated the AMI group in a 1-year patients' follow-up study. We used two different tests to detect anti-C. pneumoniae antibodies: recombinant enzyme immunoassay antilipopolysaccharide antibodies and a reference microimmunofluorescence test. RESULTS: High titers of C. pneumoniae microimmunofluorescence antibodies were found in 89.65% of the AMI group and in 25% of the HCM group (p = 0.0000065). Immunoglobulin A-microimmunofluorescence was 51.72% in the AMI group and 20.83% in the HCM group (p = 0.0042). Immunoglobulin G and immunoglobulin A antilipopolysoccharide titers were 65.51% and 62.60% in the AMI group and 20.83% in the HCM group, respectively (p = 0.006). High concentrations of interleukin-6 were found in 86.20% of our AMI group (p value = 54.38 pg/ml) when compared with the control group. A good correlation between interleukin-6 levels and immunoglobulin A-lipopolysaccharide titers (r = 0.658) was found. CONCLUSION: The presence of a high prevalence rate and high titers of immunoglobulin G and immunoglobulin A-specific anti-C. pneumoniae antibodies in AMI at admission demonstrated the presence of a specific anti-C. pneumoniae immunization in the AMI population.

Aged↗

HIV-specific mucosal and cellular immunity in HIV-seronegative partners of HIV-seropositive individuals.

HIV-specific mucosal and cellular immunity was analyzed in heterosexual couples discordant for HIV status in serum and in HIV-unexposed controls. HIV-specific IgA but not IgG was present in urine and vaginal wash samples from HIV-exposed seronegative individuals (ESN), whereas both IgA and IgG were observed in their HIV-seropositive partners; antibodies were not detected in low-risk controls. Envelope protein (Env) peptide-stimulated interleukin-2 (IL-2) production by peripheral blood mononuclear cells (PBMCs) was detected in 9 out of 16 ESNs, 5 out of 16 HIV-infected patients and 1 out of 50 controls. Env peptide-stimulated PBMCs of ESNs produced more IL-2 and less IL-10 compared with those of HIV-infected individuals; no differences were observed in chemokine production or in CCR5 expression. These data demonstrate that a compartmentalized immune response to pathogens is possible in humans and raise the possibility of protective roles for cell-mediated immunity and mucosal IgA in HIV-seronegative individuals exposed to HIV.

Adult↗

Esterase-like activity of human serum albumin toward prodrug esters of nicotinic acid.

The esterase-like activity of human serum albumin (HSA) toward esters of nicotinic acid was investigated under a variety of conditions such as protein concentration, temperature, pH, ionic strength, nature of buffers, and presence of organic solvents. Initial rate constants of hydrolysis of 18 nicotinates in the presence of 50 microM HSA were measured at pH 7.4 and 37 degrees C. The substrates displayed half-lifes ranging from less than 15 min (2-butoxyethyl nicotinate) to more than 95 hr (methyl nicotinate). The hydrolysis of tert-butyl nicotinate was too slow to be measurable, whereas 1-carbamoylethyl nicotinate was stabilized against hydrolysis by the presence of HSA. The rate constants of HSA-catalyzed hydrolysis were well correlated (r2 = 0.85; N = 12) with previously published data obtained in human plasma, indicating similar substrate specificities in the two biological preparations. All evidence points to serum albumin as the possible major catalyst of hydrolysis of nicotinate esters in human plasma.

Esterases↗

[Anti-Chlamydia pneumoniae antibodies and production of interleukin 6 in acute myocardial infarct].

UNLABELLED: Chlamydia pneumoniae (C.p.) has been correlated with acute myocardial infarction (AMI). High levels of anti-C.p. antibodies and circulating immune complexes containing C.p. lypopolyaaccharide (LPS) antigens have been demonstrated in AMI. LPS antigen and especially Chlamydial LPS is one of the best antigen and it is also a very good Interleukin inductor. Moreover, interleukin 6 (IL-6) has been observed in AMI patients. The aim of our study was to assess the possible relationships between anti-C.p. immune response and IL-6 production in AMI patients. We studied 17 consecutive patients with myocardial infarction (12 males and 5 females; mean age 62; range 46-72). Blood samples were obtained immediately after hospital admission. There were 17 control subjects (HCM) (mean age 62; range 45-72) who were matched for the main coronary risk factors (gender, age, diabetes, hypertension, hypercolesterolemia, smoking, family history of ischemic heart disease). In addition, we evaluated the AMI patients in a one-year follow-up study (FU). RESULTS: High levels of C.p. IgG MIF were found in 82.3% of our AMI patients and in 29.4% of HCM subjects (p = 0.0000065). IgA-MIF were 70.5% in AMI patients and 29.4% in HCM (p = 0.0042). High levels of C.p. IgG and IgA anti-LPS were found, with a very high prevalence rate of 76.4% and 64.7% in AMI patients, and both rates were 47.0% (p = 0.158; p = 0.489) in HCM. Very high levels of IL-6 were found (m = 54.38 pg/ml) in 100% of the AMI patients (normal values in our population: 0-10.86 pg/ml) and only detectable levels in 5.8% of HCM. A good linear correlation was demonstrated between IL-6 and IgA levels in the first sample (r = 0.655). The high levels of anti-C.p. IgG, IgA and IL-6, with a good correlation between IL-6 and IgA levels, may confirm the presence of an active infection and probably of a reinfection.

Acute Disease↗

[Effectiveness of the Palmaz Schatz stent in elderly patients with unstable angina: factors predictive of short-term and long-term events].

INTRODUCTION: To evaluate clinical, procedural and therapeutical predictors of in- and out-hospital events in the elderly, we analyzed 69 consecutive patients (age: 74, range: 70-87) with unstable angina who successfully underwent Palmaz Schatz coronary stenting. METHODS: Between March 1991 and March 1994, after the stenting procedure, a cohort of 38 patients (AC) was treated with iv heparin for 48 hours, warfarin (dosage titrated on INR) and dipiridamole (75 mg tid) for 3 months, aspirin (325 mg a day) chronically; between April 1994 and April 1995, after 48 hours of iv heparin, a second cohort of 31 patients (NO AC) received subcutaneous low molecular weight heparin (4000 U a day) for a week, ticlopidine (250 bid) for 1 month, and aspirin (100 mg a day) chronically. NO AC patients showed, by protocol, a higher postdilatation pressure (14 +/- 2 vs 9 +/- 3 atm, p < 0.0001). RESULTS: The 2 cohorts of patients were similar with respect to baseline clinical and angiographic findings. A shorter hospital stay (5 +/- 2 vs 10 +/- 6 days, p < 0.0001) and a lower incidence of in-hospital events were seen in the NO AC group (3.2 vs 24%, p = 0.028), both by a reduction of ischaemic events (3.2 vs 10.5%, p = ns) and hemorrhagic events (0 vs 13.2%, p = 0.03). During a mean follow-up of 21 +/- 13 months, NO AC patients did not show a significant lower rate of out-hospital events (1 year event-free survival respectively 94.7% in NO AC cohort vs 85.7% in AC cohort, p = ns). At logistic regression model, anticoagulant therapy (OR 10.89, Cl 1.39-85.28, p < 0.05) and refractory angina (Braunwald C3) (OR 5.70, Cl 1.12-29.03, p < 0.05) were significantly related to the incidence of acute events, while refractory angina (OR 5.76, Cl 1.27-26.00, p = 0.02) and multivessel disease (OR 3.31, Cl 0.89-12.20, p = 0.07) to the occurrence of late cardiac events, particularly for a higher risk of non-target site new revascularizations. Stent implantation on saphenous vein graft was also associated to a higher risk of repeating a revascularization of non-treated sites (20 vs 4%, p = 0.021). CONCLUSIONS: In elderly with unstable angina treated with Palmaz Schatz stenting, NO AC patients showed a significant reduction of in-hospital events without a subsequent higher risk of late events. In addition, refractory angina, multivessel disease and stent implantation on saphenous vein graft were the other main clinical variables predictive of out-hospital events particularly for higher risk of non-target site new revascularizations.

Age Factors↗

[Clinical polymorphic presentation and natural history of active myocarditis: experience in 60 cases].

Eight-hundred thirty patients (pts) with suspected myocardial disease of undefined etiology were observed from 1978 to 1996. In 350 pts, the clinical diagnosis was of dilated cardiomyopathy (DCM) or myocarditis. An endomyocardial biopsy was performed on all patients and in 54 of them (15%), an active myocarditis was identified. In six cases, myocarditis was detected at autopsy. There were 37 male patients and 23 females, with an average age of 35.5 +/- 15 years (range 1.67). Mean time interval between clinical onset and diagnosis was 4 +/- 10 months. Clinical presentation was characterized in 4 cases by fulminant myocarditis (Group I), in 8 cases by chest pain (Group II), in 14 cases by arrhythmia (Group III: hypokinetic in 9 pts and hyperkinetic in 5) and, in the last 34 pts, by congestive heart failure (CHF) (Group IV). Improvement was defined at 9 +/- 3 months according to a clinical score based on left ventricular shortening fraction (increase > or = 5 units), New York Heart Association Class improvement by (at least one Class) and left ventricular end-diastolic diameter (decrease > or = 10%). The main clinical and instrumental parameters characterizing the groups were: a more severe dilatation and left ventricular dysfunction in the pts belonging to Group I or IV with respect to those in Group II and III; a significantly worse prognosis in terms of evolution in DCM or death/cardiac transplantation (CT) in the pts from the Group II and III. After a follow-up period of 48 +/- 46 months, the mortality in the four groups was: 100% (4/4), 0% (0/8), 21% (3/14), 38% (13/34). Fifty percent of deaths were concentrated in the first 2 years of follow-up. Left ventricular end-diastolic diameter (OR 1.09, p < 0.05), age (OR 0.95), presence of left ventricular bundle branch block (OR 2.32), right ventricular function (OR 2.43) at clinical onset and the status of improvement at 9 +/- 3 months of follow-up (OR 0.24, p < 0.05) are predictors of evolution in DCM or death/CT for the pts with onset from CHF (Group IV). Immunosuppressive treatment has been utilized for the 76% of the pts. No conclusion can be drawn on the efficacy of this therapy, but no adverse events significantly related to therapy have been observed in a 9 +/- 3 months follow-up period. In conclusion, myocarditis can show a clinical presentation polymorphism, which influences the prognosis and natural history of the disease. Evolution in DCM and adverse events (death/CT) are more common in Groups I and IV. Some simple parameters evaluated at clinical presentation and the proposed classification as "improved" or "not improved" after a short-term follow-up (9 +/- 3 months) show good predictive accuracy. The present study does not allow us to draw any conclusion about the efficacy of immunosuppressive treatment. A randomized, controlled, large-scale trial, with adequate follow-up and advanced histological diagnosis techniques will help define the role of immunosuppressive therapy and patient eligibility criteria for this treatment.

Acute Disease↗

A continuous fluorimetric method to monitor the enzymatic hydrolysis of medicinal esters.

This paper describes a new spectrofluorimetric assay for continuously monitoring the enzymatic hydrolysis of medicinal esters. The procedure is based on the stoichiometric quantity of protons generated by the hydrolysis of the substrate, which produces changes in the fluorescence of a pH-sensitive dye. The pH indicator, 2', 7'-bis(car-boxyethyl)-5(6)-carboxyfluorescein, was selected due to its favourable pKa for studies under physiological conditions. Moreover, the presence of a domain in the spectra (< 442 nm) where fluorescence intensities are independent of pH allows measurements of wavelength ratios that cancel artifacts and lower sample-to-sample variability. The indicator did not affect the catalytic activity of purified hog liver carboxylesterase or human serum albumin. This assay is easy to perform and appears to be especially useful for studying enzymatic reactions with half-lives of the order of minutes or hours.

Animals↗