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Biomedical subjects

E Boschetti

Publications and source records attributed to E Boschetti.

At least 37 records · Page 2Linked to original sources
PubMed1993

Features of thrombi and diagnostic accuracy of impedance plethysmography in symptomatic and asymptomatic deep vein thrombosis.

Impedance plethysmography (IPG) has high sensitivity and specificity in patients with symptomatic deep vein thrombosis (DVT) while it fails to detect asymptomatic DVT. The aim of this study was to determine whether the features of thrombi such as location, size and occlusiveness could explain the different accuracy of IPG in symptomatic and asymptomatic DVT patients. One-hundred and seventeen consecutive outpatients with a clinical suspicion of DVT and 246 consecutive patients undergoing hip surgery were admitted to the study. In symptomatic patients IPG was performed on the day of referral, followed by venography, while in asymptomatic patients IPG was performed as a surveillance programme, followed by bilateral venography. A venography proved DVT was observed in 37% of the symptomatic patients and 34% of the asymptomatic limbs. A significantly higher proportion of proximal DVTs was found in symptomatic patients than in asymptomatic patients (78% vs 46%; p = 0.001). The mean Marder score, taken as an index of thrombus size, was significantly higher in symptomatic patients than in asymptomatic patients (19.0 vs 9.6; p = 0.0001). A significantly higher proportion of occlusive DVTs was observed in symptomatic than in asymptomatic patients (69% vs 36%; p = 0.001). We conclude that the unsatisfactory diagnostic accuracy of IPG in asymptomatic DVT is due to the high prevalence of distal, small and non occlusive thrombi. Such thrombi are unlikely to cause a critical obstruction of the venous outflow and therefore to produce a positive IPG.

Dermatan Sulfate↗
PubMed1993

[Chromatographic sorbents for the preparative separation of proteins].

Chromatography separation steps are technologically very well accepted in downstream bioprocessing, but progresses are expected on productivity level as well as on cleanability. At large production scale, the driving force for purification processes is the throughput intended as the amount of material separated at a given velocity and for a defined level of purity. The possibility to play on some phases of the separation cycle and on the variation of dynamic binding capacity are clearly useful to diminish the separation time while maintaining high levels of loads. On the other hand, to sanitize properly the sorbents, it has been found that only very strong solutions such as alkaline ethanol or alternated acidic-alkaline solutions are effective. In this general situation, special sorbents are now proposed, withstanding stringent operational situations.

Chromatography, Liquid↗
PubMed1992

Immunoenzymatic assay of dyes currently used in affinity chromatography for protein purification.

Cibacron Blue F3-GA, Basilen Blue E3-G and Procion Red HE-3B are dyes currently used in affinity purification, and are commonly determined by spectrophotometry with limited sensitivity. An assay method is described based on a specific immunochemical recognition of the dyes amplified by a final enzymatic reaction. The sensitivity is close to 1 ng/ml of dye and the method is applicable any time that sensitive and accurate results are necessary. This method has actually been applied with success to the determination of trace amounts of dyes in the presence of affinant protein. The method was also applied to the demonstration of dye leaching from affinity sorbents when treated under acidic and/or alkaline conditions.

Adsorption↗
PubMed1992

Immunochemical quantification of procion red HE-3B used as ligand in affinity chromatography.

The quantification of Procion Red HE-3B used as a ligand in affinity chromatography for proteins is reported. It's based on an enzyme-linked immunosorbent assay using antibodies against the dye. Polyclonal antibodies were classically prepared after conjugation of the dye on KLH and injection into rabbits. The development of the assay was based on the competitive inhibition between hemoglobin-dye complex and free dye. The sensitivity of this method was about 1000-times higher than a classical spectrophotometric assay, and was modulated by some chemical substituents attached on the native dye. It was demonstrated that the assay was applicable to the determination of dye traces that may be released from dye affinity sorbents. Moreover, the quantification of the dye was successfully applied to proteins that are being purified from a dye affinity column.

Chromatography, Affinity↗
PubMed1992

A sensitive enzyme immunoassay for the detection of a synthetic affinity ligand, the reactive yellow 13 dye.

Dye-affinity chromatography is widely and increasingly used for the isolation of various proteins. In particular, the purification of transthyretin can be efficiently achieved by affinity chromatography on immobilized Reactive Yellow 13. Measurement of trace-amounts of dye leaching from affinity columns is important because of possible toxicity or side effects linked with the presence of dye in therapeutic transthyretin preparations. A competitive enzyme immunoassay was developed to monitor yellow-dye column leaching. Biotinylated rabbit anti-Reactive Yellow 13 antibodies (immunoglobulin G fraction) were used as principal reagent. The assay is specific sensitive to 1 ng/ml of Reactive Yellow 13, has a good reproducibility and allows the accurate detection of the dye in the presence of transthyretin.

Affinity Labels↗
PubMed1992

Antiemetic activity of two different high doses and schedules of metoclopramide in dacarbazine-treated cancer patients.

The antiemetic activity of two different high doses and schedules of metoclopramide in dacarbazine-treated cancer patients was compared in a double-blind crossover study. Regimen A consisted of metoclopramide [2 mg/kg x 4 intravenously (i.v.)] plus methylprednisolone (250 mg x 2 i.v.) plus diphenhydramine (50 mg x 2 i.v.). Regimen B consisted of metoclopramide (3 mg/kg x 2 i.v.) plus dexamethasone (20 mg i.v.) and diphenhydramine (50 mg i.v.). Both treatments were administered for the first 2 days of 5-day dacarbazine chemotherapy. Thirty-two patients (13 men and 19 women) affected by melanoma and sarcoma were entered in the study. Complete protection against nausea and vomiting for the first 2 days of chemotherapy in both antiemetic regimens was not significantly different. Patient preference and tolerance of the two antiemetic treatments were similar. Regimen B, employing a lower dosage of metoclopramide and steroids and using a more simple schedule of administration should be the preferred treatment.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1992

ELISA-based quantification of cibacron blue F3GA used as ligand in affinity chromatography.

The development of an enzyme-linked immunosorbent assay (ELISA) for Cibacron Blue F3GA is reported. It quantifies a trace amount of blue dye used as an affinity chromatography ligand, rendering possible the measurement of traces of leached dye. Polyclonal antibodies were prepared after conjugation on the dye on KLH and injection into rabbits. The ELISA test was based on the competitive inhibition between a hemoglobin-dye complex adsorbed on the wells of a titration microplate and a free dye. The sensitivity of the assay was about 1000 times higher than a classical spectrophotometric assay and was modulated by some chemical substituents to the dye. The described ELISA assay was also successfully applied to the quantification of dye traces in the presence of human albumin.

Adsorption↗
PubMed1992

[Immunoenzymatic assay of dyes used in affinity chromatography of proteins].

Immobilized reactive dyes are frequently used in pseudoaffinity chromatography since it has been evidenced that they could specifically interact with some biologicals of interest, and, as a consequence, be separated and purified by this way. This specificity was so high in some cases, that a single step purification process was possible. In addition to these features, the wide variety of dye molecules available associated to their chemical stability and low cost make possible their use as ligands for industrial applications. Nevertheless, one drawback of the use of dye sorbents for protein purification is the possible leakage of dye molecules, with a risk of biological properties alteration of the purified product. We describe a new quantitative assay method for dyes that could contaminate biologicals separated through special affinity columns. This method based on the specific recognition of dyes by antibodies showed a high specificity and a much better sensitivity than the classical spectrophotometric approaches. It has been applied to the quantification of dyes used in affinity chromatography (Cibacron Blue F3-GA, Basilen Bleue E3-G and Procion Red HE-3B) in the presence of proteins having a special affinity for these dyes. This immunoenzymatic assay is also easily applicable to the detection of leachables from dye affinity columns submitted to drastic regeneration treatments.

Animals↗
PubMed1991

Immobilized cibacron blue--leachables, support stability and toxicity on cultured cells.

Although immobilized dyes are widely used on the laboratory scale and have good potential for industrial applications, they are still subject to some reservations. Little information is available about dye leakage and toxicity, which seriously hinders the use of such supports in the production of pure proteins. Investigations of the leakage mechanism and the in vitro toxicity of the native dye and of that leached from the column are reported. The possible presence of traces of dye in the purified biological materials necessitates the availability of sensitive analytical tests. The preparation and preliminary isolation of dye antibodies as a first step in the development of an immunohistochemical assay of leached dyes are also described.

Antibodies↗
PubMed1991

Double-blind crossover trial of single vs. divided dose of metoclopramide in a combined regimen for treatment of cisplatin-induced emesis.

In a double-blind crossover antiemetic study in cisplatin-treated cancer patients, metoclopramide 4 mg/kg as a single intravenous dose (regimen A) was compared with 3 mg/kg in two doses (regimen B). In both regimens, metoclopramide was combined with dexamethasone and diphenhydramine. 65 consecutive, chemotherapy-naïve inpatients (45 males and 20 females) treated with high doses (at least 50 mg/m2) of cisplatin entered the study and 54 completed both treatments. Complete protection from vomiting and nausea, mean number of emetic episodes, mean maximum intensity of nausea and mean duration of emesis or nausea were similar with the two antiemetic regimens. 23 patients (43%) did not express a treatment preference, while 16 (30%) preferred regimen B and 15 (28%) preferred regimen A. Side-effects were similar with the two metoclopramide schedules. A combined antiemetic regimen of a single high dose of metoclopramide (4 mg/kg) can preserve efficacy and tolerability and thus should be preferred.

Cisplatin↗
PubMed1991

Intra and interobserver variability in cancer patients' performance status assessed according to Karnofsky and ECOG scales.

The Karnofsky (K) and ECOG (E) performance status (PS) scales are widely used to evaluate the functional status of cancer patients to determine their eligibility for clinical trials and their prognosis, but knowledge of inter and intraobserver variability of these scales is scarse. We therefore planned a prospective study on 209 consecutive cancer patients to evaluate this critical point. Two independent observers evaluated the KPS and EPS of each patient by interviewing them on the same day. After their interviews the patients were asked to fill in, again on the same day, a self-evaluation scale concerning their ability to perform the routine activities of daily life. The 209 patient self-evaluation scales were presented twice, randomly and blinded, to the two observers who had participated in the evaluation of PS as well as to one other observer who had not. The interobserver correlation for both scales was very high (K = 0.921 for KPS and K = 0.914 for EPS) as was the intraobserver correlation (for KPS: K = 0.993, K = 0.960, and K = 0.959 and, respectively, for EPS: K = 0.982, K = 0.970, and K = 0.920). On the basis of these results, it appears that evaluation of PS made by a clinical oncologist using K or E scales can be very reliable and is a guarantee of optimal selection of cancer patients for inclusion in clinical trials.

Aged↗
PubMed1991

Predictive factors of delayed emesis in cisplatin-treated patients and antiemetic activity and tolerability of metoclopramide or dexamethasone. A randomized single-blind study.

To prevent delayed emesis induced by cisplatin (mean dose 90 mg/m2), 120 consecutive patients were randomized to receive, in a 7-day crossover design, oral metoclopramide (20 mg q.i.d.), dexamethasone (1 mg q.i.d.) or placebo (two tablets q.i.d.) starting 24 hours after the end of chemotherapy. Complete protection from nausea, but not from vomiting. was significantly increased by both dexamethasone and metoclopramide with respect to placebo. Important prognostic factors favoring the appearance of delayed emesis were incomplete protection from vomiting during the first 24 hours after cisplatin, female gender, and highest cisplatin doses. Tolerability of both drugs was good. Larger and randomized controlled trials are necessary to identify better preventive treatment of delayed emesis induced by cisplatin.

Cisplatin↗
PubMed1991

Large scale production of human albumin: three years experience of an affinity chromatography process.

Immobilized Cibacron Blue is a well-known tool for the separation of a number of proteins and enzymes. Among them, human plasma albumin was easily purified from crude plasma and consequently, this approach represents an interesting way for a production scale. Our data describes about three years of experience in the production of human albumin using large columns of immobilized Cibacron Blue. The amount of albumin produced per cycle was about 250 g on a column of about 50 l. Over 500 cycles the final purity of albumin was very high and constant (98-100%). The albumin yield of the chromatographic fractionation was approximately 82%. The column performance remained acceptable when regeneration operations were applied to the sorbent. The long life of the sorbent renders this approach very attractive and economically acceptable for large scale applications.

Chromatography, Affinity↗
PubMed1991

[Defined media for hybridoma culture: do they really replace serum and why are they attractive?].

The cell culture efficiency of serum free media is still the main question which limits the use of such a product. However, the experience due to a long usage of serum free media allows to certify that for hybridomas the proliferation growth is often similar to the levels attained with serum. Antibodies secretion is also good or superior to the one obtained with classical media used in different culture devices like flasks and cytocultures. Finally the higher purity degree of monoclonal antibodies in the cell culture supernatant is also a major advantage of serum free media.

Animals↗
PubMed1990

Composite affinity sorbents and their cleaning in place.

Making large-scale affinity sorbents that are reusable under acceptable hygienic conditions implies specific treatments for cleaning in place with known aqueous solutions of chemical agents. However, common agents such as sodium hydroxide are frequently considered too drastic for the stability of macromolecular biologically active immobilized ligands. According to a large series of trials, it was found that only a mixture of sodium hydroxide and ethanol was actually effective in sterilizing a sorbent in a single step. When hydroxide or an ethanol-acetic acid mixture were used alone, they were not totally efficient in the inactivation of sporulated Bacillus subtilis. Conversely, they were efficient when used sequentially. All these solutions were able to remove pyrogens from chromatographic sorbents. As the sterilizing solutions contained a certain amount of ethanol, the most suitable chromatographic affinity sorbents had to be based on an incompressible matrix. When washing an affinity silica sorbent that had proteins as ligands with solutions such as sodium hydroxide, ethanol-acetic acid or ethanol-sodium hydroxide, it was found that certain sorbents were able to tolerate the treatments without a noticeable decrease in their biochemical activity.

Acetates↗
PubMed1990

Effect of aspirin and dipyridamole treatment on prostacyclin production by human veins.

Patients admitted for surgical removal of varicose veins were treated in a blinded manner for 48 hours prior to surgery with either placebo, low-dose aspirin (25 mg twice daily), dipyridamole (150 mg twice daily) or both. Segments of vein excised at surgery were incubated with or without sodium arachidonate and subsequent prostacyclin (PGI2) production was measured without knowledge of treatment given. During the first 5 minute period of incubation in the presence of arachidonate, veins from dipyridamole-treated patients demonstrated increased (by 75%) arachidonate-stimulated PGI2 production compared to placebo-treated patients. By contrast, PGI2 production was reduced by 64% by aspirin treatment and 67% by aspirin plus dipyridamole compared to placebo-treated patients (p = less than 0.05). In unstimulated vein segments incubated in the absence of arachidonate, spontaneous PGI2 production during the first 5 minute incubation period was increased 32% following dipyridamole treatment but was unchanged following aspirin treatment. By contrast, unstimulated (spontaneous) PGI2 production in patients treated with aspirin plus dipyridamole was reduced by 57% (p = less than 0.05), compared to both placebo- and aspirin-treated patients, and by 71% (p = less than 0.05) compared to dipyridamole-treated patients. With repeated change of incubation medium, the ability of vein walls to produce PGI2 declined. This exhaustion was not prevented by drug treatment. However, drug effects between patient treatment groups were consistent over successive incubation periods. These results suggest that certain therapeutic benefits that might be achieved by enhancement of PGI2 production from vascular endothelium following dipyridamole treatment may be reduced by simultaneous aspirin treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Aspirin↗