提高PLGA微球载蛋白多肽类药物稳定性添加剂的研究进展

聚乳酸-羟基乙酸共聚物(PLGA)包裹药物制成微球制剂是近年来药物输送体系的研究热点,但是PLGA包囊药物尤其是蛋白多肽类变性失活问题是研究的难点.改进生产工艺特别是使用添加剂可提高微球中包载药物的稳定性.综述了PLGA微球制备及释放过程中不同添加剂的作用特别是稳定蛋白药物的作用.     

Angiostatic proteins and peptides

Angiogenesis, or the formation of new vasculature out of preexisting capillaries, is a sequence of events that is essential in the normal physiological processes of tissue growth and in a broad spectrum of pathologies. The diseases in which angiogenesis plays a key role are divided into diseases that are characterized by hypoxia/ ischemia and diseases that are dependent on neovascularization. The formerpathologies may benefit from therapeutic angiogenesis stimulation. This review concentrates on the different strategies to inhibit angiogenesis in diseases that are characterized by excessive angiogenesis, for example, cancer, arthritis, diabetic retinopathy, and inflammatory diseases. These diseases are dependent on the development of newvasculature, and hence, a large variety of different strategies to inhibit angiogenesis are underwayin laboratories throughout the world. At present, over250 angiogenesis inhibitors are described, and approximately half of them display activity in in vivo models. A large percentage of these molecules are natural, nonnatural, or synthetic so-called small molecules. Others are of protein origin, either endogenous or exogenous by nature. The authors highlight the current knowledge on the development of angiostatic proteins and peptides and their potential in the treatment of disease.     

Selected pool of peptides from ESAT-6 and CFP-10 proteins for detection of Mycobacterium tuberculosis infection

We have validated a new test for detecting Mycobacterium tuberculosis infection. A pool of synthetic peptides derived from ESAT-6 and CFP-10 proteins was used to detect the number of specific gamma interferon-producing T cells by means of an enzyme-linked immunospot assay. Sixty-eight individuals positive for M. tuberculosis infection, either human immunodeficiency virus-seropositive or -seronegative, were studied. The test results were highly specific (87.5%) and sensitive (93.1%), more so than a classical lymphoproliferative assay (specificity and sensitivity of 77.27%), opening new possibilities for diagnosis and screening of tuberculosis. Moreover, the test allowed us to distinguish individuals infected with M. tuberculosis from those vaccinated with BCG.     

Renal handling of proteins and peptides

The kidney plays an important role in the metabolism of proteins and peptides. Current evidence indicates that only the proximal tubule possesses the mechanisms for absorption, transport and/or degradation of these substances. Large proteins and polypeptide molecules filtered by the glomerulus, are absorbed from proximal tubular fluid by luminal endocytosis into apical vacuoles which fuse with primary lysosomes where hydrolysis occurs followed by diffusion of metabolites out of the cells and into the blood. Recent evidence indicates that small peptides are handled by a different mechanism. It appears that small peptides are degraded at the luminal surface of the brush-border of proximal tubules, which contains many hydrolytic enzymes, by the process of membrane or contact digestion with reabsorption of the breakdown products. Proximal tubular mechanisms for handling of proteins and peptides are probably important biologically to conserve amino acids, inactive toxic substances and help regulate the circulating level of protein and peptide hormones.     

Molecular techniques in mycobacterial detection

OBJECTIVE: To assess the clinical utility of the commercial nucleic acid amplification (NAA) tests (ie, Amplified Mycobacterium Tuberculosis Direct Test, Gen-Probe, Inc and AMPLICOR Mycobacterium tuberculosis Test, Roche Molecular Systems, Inc) for direct detection of Mycobacterium tuberculosis complex. DATA SOURCES: Review of the English-language literature. CONCLUSIONS: The performance of both NAA tests is excellent (sensitivity, > or = 95%; specificity, 100%) when testing respiratory specimens that are smear-positive for acid-fast bacilli (AFB). Only the Gen-Probe assay is approved for testing respiratory specimens regardless of the AFB smear result. Data from 3 studies showed that the sensitivity of the Mycobacterium Tuberculosis Direct Test in smear-negative patients ranged from 83% to 85%, and that the specificity was 99%. Both NAA tests have been used to test nonrespiratory specimens; in some studies, the performance was comparable to the performance obtained for respiratory specimens, whereas in others, it was lower. The NAA tests also appear to be reliable tools for rapid detection of M tuberculosis complex in positive broth cultures of all specimen types (except blood). The impact of the NAA tests on patient outcome varies based on the result of the AFB smear. In smear-positive patients, public health and hospital infection-control resources are predominantly affected. The potential for influencing patient outcome is much greater when the AFB smear is negative. In smear-negative patients, the NAA test could provide more rapid diagnosis of tuberculosis and subsequent initiation of therapy; eliminate the need for invasive diagnostic procedures, which are both costly and pose an added risk to the patient; and allow earlier discharge of hospitalized patients. Prospective studies concerning the cost-effectiveness of the NAA tests are needed.     

兴奋剂检测技术及其进展

本文介绍了兴奋剂检测技术的发展概况,阐述了应用于兴奋剂检测样品的前处理,分离及分析方法,并探讨了在线提取柱切换检测技术及灵敏,快速的高通量检测技术等的应用前景.     

Separation and characterization of cardiac antigen proteins

An attempt has been made to purify and characterize the protein antigens related to autoimmune carditis in the heart extract of albino rats by ammonium sulphate fractionation, molecular sieve gel filtration and polyacrylamide gel electrophoresis. The heart extract of albino rat at pH 7·0 contains at least nine proteins out of which three are antigenic in rabbits and have been termed as CA I, II and III. These antigens can be purified by ammonium sulphate fractional precipitation followed by Sephadex G-100 gel filtration. Their molecular weights were 6·6 (±2) × 10⁴, 2·8 (±0·85) × 10⁴, 4·8 (±1·5) × 10⁴ Daltons. A fourth antigen (CA IV) detected is probably polysaccharide.     

Serological techniques for detection of antibody to galactocerebroside

Two serological techniques were developed for the detection of antibody to galactocerebroside using liposomes as a carrier for the lipid hapten. One assay is a radioimmunoprecipitation test employing [³H]cholesterol as a marker in the galactocerebroside-liposomes. The other is a less sensitive but quick and easy galactocerebroside-liposome agglutination assay.Specificity is demonstrated by comparison of titers when other lipid haptens replace galactocerebroside in the liposomes, and when other anti-glycolipid antisera are reacted with galactocerebroside-liposomes.     

Flow cytometric techniques for the detection of microorganisms

Flow cytometry (FCM) is a technique, which allows one to analyse cells rapidly and individually, and permits the quantitative analysis of distributions of a property or properties in a population. It therefore offers many advantages over conventional measurements for the analysis of biological cells. Historically the technique has been widely applied for the study of mammalian cells, but its use in microbiology has been more limited; this is mainly a consequence of the smaller size of microbes, which results in the smaller optical signals that can be obtained from them. Developments in light sources and optics, together with brighter, spectrally-diverse dyes have reduced this barrier over recent years and the flow cytometer is now an essential tool in many microbiological research establishments. FCM has an increasing role to play in the detection of microbes in both industrial and clinical settings. Environmental monitoring to prevent outbreaks of human diseases such as cryptosporidiosis and Legionnaires' disease and to detect acts of biowarfare or bioterrorism are all amenable to flow cytometric study. This review seeks to highlight the role of the flow cytometer in the detection of microbial cells.     

Temperature and pH-responsive polymeric composite membranes for controlled delivery of proteins and peptides

This work was focused on the investigation of temperature and pH-responsive polymeric composite membranes and their permeability to proteins and peptides in response to environmental stimuli. The composite membranes were prepared from nanoparticles of poly(N-isopropylacrylamide-co-methacrylic acid) of various NIPAAm:MAA ratios dispersed in a matrix of a hydrophobic polymer. N-Benzoyl-L-tyrosine ethyl ester HCl, momany peptide, Leuprolide, vitamin B(12), insulin, and lysozyme were used as model solutes. The morphology of the membranes was examined with SEM and permeation of the solutes was measured using side-by-side diffusion cells at varied temperatures and pH. Permeability of the solutes across the membranes increased with increasing temperature or particle concentration, while decreased with increasing pH and molecular size of the solutes. Membranes containing nanoparticles of more NIPAAm units exhibited higher thermal sensitivity, and those with higher MAA content showed more pH responsiveness, which was in line with the temperature and pH-responsive volume change of the nanoparticles. The change in permeability was quickly detected following the application of the stimuli. These results and partition study using vitamin B(12) supported the proposed gel-pore mechanism of solute permeation through these composite membranes.     

Conjugation of synthetic peptides to carrier proteins for cell adhesion studies

Summary The ability to reproduce active site sequences from integrin receptor ligands as synthetic peptides has revolutionized the field of cell adhesion. Most studies with these peptides use them in soluble form as inhibitors of integrin function, however, other studies necessitate immobilization of the peptides onto plastic or glass surfaces for analyzing the promotion of cell adhesion. Unfortunately, peptides rarely adsorb well and adsorbed peptides may have their active residues masked by the substrate. An alternative approach is therefore to analyze peptide activity after covalent conjugation to an inert carrier protein. Methods for the production and characterization of peptide conjugates are described.     

Serological techniques for detection of citrus tristeza virus.

Citrus tristeza virus (CTV) is the most economically important virus disease of citrus. In the last ten years, remarkable progress has been achieved in the development and improvement of new serological methods for CTV detection so that serology has become a dependable tool for many research, extension and regulatory purposes worldwide. CTV-specific polyclonal antisera and monoclonal antibodies have been developed in different research laboratories and used extensively in a wide range of different studies. This review describes the diverse serological methods developed for CTV detection and analyzes the advantages, disadvantages, relative sensitivity, applications, and present status of each method.     

Molecular techniques for the detection of Chlamydia trachomatis.

A DNA probe assay (PACE; Gen-Probe, San Diego, Calif.) was compared with a culture reference method for the detection of Chlamydia trachomatis. Using stock isolates of each of the 15 serovars (A to K, Ba, L1, L2, and L3) of C. trachomatis, the lower limit of sensitivity for the DNA probe ranged between 1,086 inclusion-forming units (IFU) for serovar E (Bour) to 2,930 IFU for serovar L1 (440), with the only exception being serovar C (TW-3), with which 99 IFU was detected. There was no cross-reactivity with Chlamydia psittaci (Texas turkey) and Chlamydia pneumoniae (TWAR-183). Bacterial and fungal isolates representing 14 species of normal vaginal flora as well as Neisseria gonorrhoeae gave negative results with the DNA probe when tested at a level of 1.5 X 10(7) CFU/ml. In addition, the DNA probe, a direct fluorescent-antibody stain (DFA) (MicroTrak; Syva Corp., Palo Alto, Calif.), and an enzyme-linked immunosorbent assay (Chlamydiazyme; Abbott Laboratories, North Chicago, Ill.) were compared with culture for the detection of C. trachomatis, using 196 clinical cervical samples. Of the 196 samples, 20 (10%) were culture positive. Of the 176 culture-negative samples, 1 was not evaluated by DNA probe and 4, because of a lack of cellular material, were not evaluated by DFA. The sensitivities of the DNA probe, DFA, and enzyme-linked immunosorbent assay were 60, 75, and 85%, respectively, and specificities were 95, 99, and 97%, respectively. Of the false-positive direct results, there was only one specimen with which more than one direct method was positive, and with this specimen all three direct methods were positive. The majority of false-negative results by the direct methods were from specimens which by the culture method gave <100 IFU per culture.     

MLPA and MAPH: new techniques for detection of gene deletions

Screening for deletions of all or part of genes poses a challenge in the diagnostic laboratory. Numerous methods are available for detecting deletions of a few base pairs or very large deletions, but difficulties arise in detecting deletions of a few kilobases. Two new techniques have recently been described that allow detection of such mid-size deletions by simultaneously screening for the loss or duplication of up to 40 target sequences. These are the multiplex amplification and probe hybridization (MAPH) and the multiplex ligation-dependent probe amplification (MLPA). Both rely on sequence-specific probe hybridization to genomic DNA, followed by amplification of the hybridized probe, and semi-quantitative analysis of the resulting PCR products. The relative peak heights or band intensities from each target indicate their initial concentration. The two techniques differ in the ease with which probes can be generated in house, and the labor intensity of performing the assay.