非浓缩尿琼脂糖蛋白电泳及其临床应用

目的采用浓缩尿蛋白电泳,对尿蛋白组成成分进行测定,以判断肾脏疾病的受损部位及病变程度.方法采用法国SEBIA MG-300半自动电泳分析仪对88例肾脏病患儿进行尿蛋白组分分析.结果 88例尿蛋白电泳显示50例为肾小球性蛋白尿,25例为生理性蛋白尿,12例为混合性蛋白尿,1例为肾小管性蛋白尿.其中41例同时作肾穿刺术,3例肾小球轻微病变,26例局灶性肾小球肾炎,12例局灶节段性肾小球硬化.结论非浓缩尿蛋白电泳技术方法简便,无创伤性,且尿蛋白组分和肾脏病理损害程度有关,值得临床应用.     

非浓缩尿蛋白组分免疫比浊定量检测的临床应用

目的:测定尿高、中、低分子蛋白,区分尿蛋白类型并应用于临床.方法:用免疫比浊法定量检测患者尿蛋白,与肾活检病理结果对照分析.结果:将蛋白尿归纳为白蛋白为主蛋白尿、非选择性肾小球蛋白尿、混合性蛋白尿、肾小管性蛋白尿、溢出性蛋白尿5种类型,对肾脏病的诊断与肾活检病理结果相符合.尿蛋白定性阴性的糖尿病患者尿液中单独α1-微球蛋白升高率明显高于单独白蛋白升高率.结论:本法能客观反映肾脏损伤部位、程度,对肾脏病的诊断、鉴别诊断具有一定价值.     

尿蛋白电泳在小儿肾病诊断中的应用价值

目的:探讨尿蛋白电泳在小儿肾病诊断中的应用价值.方法:应用十二烷基磺酸钠-琼脂糖凝胶(SDSAGE)对642例肾病患儿24 h尿液或晨尿进行尿蛋白电泳分析,统计其尿蛋白电泳结果的构成比,并对其中54例有同时进行肾病理活检确诊的病例进行比较,计算两者间的符合率.结果:根据尿蛋白电泳图谱,蛋白尿可区分为肾小球性蛋白尿、肾小管性蛋白尿、混合性蛋白尿、单纯白蛋白性蛋白尿及未检出蛋白尿,其构成比分别为67.9%、0.9%、15.0%、11.4%和4.8%.其中54例有同时进行尿蛋白电泳及肾病理活检确诊者,两者间的符合率为94.4%.结论:非浓缩尿蛋白电泳法敏感性好、快捷简便,并能通过SDS-AGG尿蛋白电泳图谱,提示小儿肾病的病变部位及损害程度,与肾病理活检结果基本一致.     

尿蛋白SDS-AGE电泳谱带分析

目的研究各种尿蛋白的SDS--AGE电泳谱带模式及其在诊断肾脏疾病中的作用.方法 收集各种肾病病人尿液进行尿蛋白SDS-AGE电泳,扫描观察谱带的特征及各蛋白组分的百分比并进行评估分析.结果肾前性蛋白尿出现低分子量蛋白带但谱带单一;肾小管损伤性蛋白尿虽出现低分子量蛋白带但谱带较复杂;肾小球损伤性蛋白尿出现单一中分子量蛋白带和中、高分子量蛋白带;肾小球肾小管同时损伤时表现为混合性蛋白谱带(大、中、小分子蛋白带);尿路感染时尿蛋白谱带较为复杂.结论 SDS-AGE尿蛋白电泳结合尿常规分析对肾脏疾病的诊断、鉴别诊断、指导治疗及判断预后很有价值.     

尿蛋白SDS-AGE电泳谱带分析

目的 研究各种尿蛋白的SDS—AGE电泳谱带模式及其在诊断肾脏疾病中的作用。方法 收集各种肾病病人尿液进行尿蛋白SDS-AGE电泳，扫描观察谱带的特征及各蛋白组分的百分比并进行评估分析。结果 肾前性蛋白尿出现低分子量蛋白带但谱带单一；肾小管损伤性蛋白尿虽出现低分子量蛋白带但谱带较复杂；肾小球损伤性蛋白尿出现单一中分子量蛋白带和中、高分子量蛋白带；肾小球肾小管同时损伤时表现为混合性蛋白谱带(大、中、小分了蛋白带)；尿路感染时尿蛋白谱带较为复杂。结论SDS-AGE尿蛋白电泳结合尿常规分析对肾脏疾病的诊断、鉴别诊断、指导治疗及判断预后很有价值。     

分析血尿原因的新指标

血尿（即尿中出现红细胞）应当引起相当的重视。首要问题之一就是搞清血尿是起因于肾脏或是肾外，换句话说，要搞清是肾性或非肾性血尿。这将决定后续的诊断与治疗方法。鉴别肾性血尿的经典方法是检查尿沉渣中的管型，以及特殊的蛋白尿。在过去的十年中，尿中红细胞形态学检查成为一项新方法，在这一方法中，高百分比的畸形红细胞提示肾性血尿。以上这些方法都各有其局限性。比如，尿中管型比较脆弱易破坏且易漏诊；红细胞形态极大地依赖于尿标本的质量。近来，一些有趣的观察提示通过测定尿中QZ一巨球蛋白与白蛋白的比值可以区别肾小球性…     

尿蛋白电泳在肾脏疾病诊断中的临床应用

目的根据蛋白质分子量不同,采用十二烷基硫酸钠琼脂糖凝胶(SDS-AGE)对尿蛋白进行电泳,以判断肾脏疾病的受损部位及病变程度。方法采用法国Sebia公司HYDRASYS全自动电泳分析仪对104例肾脏疾病患者尿液进行电泳分析。结果根据尿蛋白电泳图谱扫描测定,104例尿蛋白电泳显示10例为生理性蛋白尿,37例为肾小球性蛋白尿,12例为肾小管性蛋白尿,45例为混合性蛋白尿,其中20例同时做肾活检,系膜增生性肾小球肾炎18例,膜增生性肾小球肾炎2例,SDS-AGE尿蛋白电泳结果与肾活检结果基本相符。结论琼脂糖尿蛋白电泳技术简便快速,无创伤,对早期肾损伤的诊断,肾脏损伤部位分析,指导治疗和判断预后有较高的价值。     

肾小管性蛋白尿检验技术的新进展

1958年Butler和Flynm指出肾小球损伤和肾小管损伤的尿蛋白成分不同,因而用来区别肾小管性蛋白尿与肾小球性蛋白尿,目前据据其产生机理把肾性蛋白尿分成以上2种类型。在肾小管性蛋白尿,有很多蛋白成分排泄于尿中,用免疫电泳就能鉴定出30种左右的成分。其中量最多的是β_2微球蛋白,免疫球蛋白的轻链,溶菌酶,视网膜结合蛋白等,最近作者等分离出来的α_1微球蛋白(分子量33,000)也证明在尿中大量排出。这些成分都存在于正常人的血浆中,正常人的尿中也可见到。由此可见,肾小管性蛋白尿的LMW(低分子)蛋白的大部分不是由肾脏的异常所产生,而是因正常血浆成分在肾小管的处理异常而出现,易通过正常肾小球的LMW蛋白因肾小管重吸收功能     

十二烷基磺酸钠-琼脂糖凝胶电泳技术在蛋白尿分析中的临床应用

检验医学传统的各种尿蛋白检测方法具有相当的局限性,难以系统、全面地分析蛋白尿的类型。近年来,建立了十二烷基磺酸钠-琼脂糖凝胶(SDS-AGE)电泳技术,进行非浓缩尿蛋白电泳对尿液各种蛋白质进行分离,并对尿蛋白各组分进行扫描定量分析,可全面地判断尿蛋白的性质。该文着重介绍该技术在鉴别各种蛋白尿中的应用。     

SLE患者非浓缩尿蛋白电泳分析

目的 探讨尿蛋白电泳分析系统性红斑狼疮(SLE)在患者肾损害诊断及疗效判断中的价值.方法 采用十二烷基磺酸钠-琼脂糖凝胶电泳(SDS-AGE)检测SLE患者非浓缩尿蛋白成分.结果 42例SLE患者检出,生理性蛋白尿1例、肾小球性蛋白尿25例、肾小管性蛋白尿2例、混合性蛋白尿14例.白蛋白的百分比升高与病情轻重呈负相关(r=-0.674)P<0.01,Ig、转铁蛋白的百分比升高与病情轻重呈正相关(r=0.635,r= 0.724)P<0.O1;并且白蛋白的百分比升高与疗效之间呈正相关(r=0.526)P<0.01,Ig、转铁蛋白的百分比降低与疗效之间呈正相关(r=0.812,r= 0.725)P<0.01.结论 尿蛋白SDS-AGE电泳对SLE患者肾损诊断及预后的判断有一定临床价值.     

Routine differential diagnosis of proteinurias by capillary electrophoresis.

Capillary electrophoresis is a relatively new analytical technique that begins to have an impact on both routine and research in clinical laboratories. Recently, a fully automated system has become commercially available (Paragon CZE 2000, Beckman, USA) for the analysis of human serum proteins. Urine protein analysis, on the other hand, is currently accomplished by electrophoresis of concentrated urine specimens. The method is used to distinguish the glomerular from the tubular proteinuria and for the identification of Bence-Jones proteins. The procedure is labor-intensive and technically demanding. We developed a technique for the serum capillary electrophoresis instrument that can also be applied routinely to the differential diagnosis of proteinurias. Overriding the programmed dilution step of the instrument, we were able to distinguish different types of proteinurias without concentration of specimens with a total protein content of 150-200 mg/l as determined by sulfosalicylic acid. The different electrophoretic patterns obtained by the capillary electrophoresis system for various specimens correlated well with established techniques (Hydragel Proteinurie Kit, Sebia, France). The method is applicable for routine analysis of urinary proteins. It is reliable, less expensive and faster than the conventional methods (electrophoretic or immunonephelometric) used today for the differentiation of proteinurias, and it can be used as a quick screening test.     

Improved measurement of urinary total protein (including light-chain proteins) with a Coomassie brilliant blue G-250-sodium dodecyl sulfate reagent

The Coomassie Brilliant Blue G-250 method for urinary proteins underestimates urinary immunoglobulin light chains when albumin or pooled serum is used as the protein standard. The specific color yields of these and other proteins can be brought closer together by adding sodium dodecyl sulfate to the reagent; however, there is some loss of sensitivity. We found such a reagent to be satisfactory for assaying urinary proteins on studying 43 patients with light-chain proteinuria, 19 of whom had multiple myeloma and six possible multiple myeloma.     

微流控芯片用于分离蛋白质的影响因素及临床应用

目的建立微流控芯片电泳分离蛋白质的方法,并探讨微流控芯片电泳在分离临床尿液蛋白组份中的初步应用价值。方法①利用自制的微流控电泳分析仪和自制的石英芯片对电泳条件进行摸索,进行微流控芯片电泳分离蛋白质的方法学重复性、检测限及抗干扰实验;②利用全自动琼脂糖电泳仪与微流控芯片电泳仪对临床尿液标本进行电泳分离,对尿液中的蛋白质进行初步定性,判断尿液中蛋白质的来源。结果①在75 mmol/L硼酸盐(pH 10.5)含1%(v/v)乙胺电泳缓冲液中,进样电压500 V,15 s时电泳蛋白可得到基线分离;②在36例尿蛋白阳性患者中,用该法检测发现溢出性蛋白尿2例、选择性蛋白尿16例、非选择性蛋白尿18例,20例健康对照未检出蛋白峰。结论该法快速、简便,检测成本极低,有利于临床的诊断和预后判断,有较好的应用前景。     

Immunonephelometric quantification of specific urinary proteins versus a simple electrophoretic method for characterizing proteinuria

OBJECTIVES: The quantification of urinary proteins presenting different molecular sizes is useful in characterizing a proteinuria. We assessed the performance of an electrophoretic system, the Hydragel Urine Profile, which allows firstly the identification of proteinuria and secondly a qualitative detection of monoclonal free light chains (FLC). DESIGN AND METHODS: Initially, the proteinuria was characterized on 127 pathological urines by quantifying albumin, a1microglobulin, immunoglobulins G by immunonephelometric quantification technique and the results were compared with the protein pattern obtained by the electrophoretic method. Secondly, we assessed the sensitivity and specificity of this electrophoretic test for the detection and characterization of Bence Jones proteins. FLC were analyzed quantitatively by an immunonephelometric assay and qualitatively by the electrophoretic test in 150 urines. RESULTS: The agreement between the two methods was good with a percentage of homology for characterizing the proteinuria of 89%. For detecting a monoclonal FLC, the electrophoretic method demonstrated a lesser sensitivity but a higher specificity compared to the immunoassay. CONCLUSION: The Urine Profile kit is a reliable assay that can be used as a screening test to differentiate the type of proteinuria.     

Difference between orthostatic and march functional proteinuria by application of stress tolerance test and SDS-PAGE

We introduced minimal necessary criteria and methods for noninvasive laboratory diagnosis and follow-up of functional proteinuria in youths: stress tolerance test, determination of total urinary proteins (TP) and their separation and identification with gradient SDS-PAGE. Renal functional adaptability in conditions of complete rest, in routine daily activity and after several hours of active physical effort has been evaluated by the tolerance test. Excretory urinary proteins, as the most appropriate markers, were analyzed with noninvasive methods. Excretory TP demonstrated the quantity and dynamics of the proteins excreted during the tolerance test. Separation with gradient (4-22.5%) SDS-PAGE provided differentiation of functional proteinuria from the other orthostatisms, through the protein fractions present and the constant finding of apolipoprotein AI. The investigation comprised 19 youths with orthostatism and 20 healthy subjects without orthostatism, all between the age of 10 and 18 years. The subjects without orthostatism excreted proteins in normal limits during the stress tolerance test. SDS-PAGE of the urinary samples, obtained during the tolerance test, in five subjects significantly differed from that in the other 14 subjects. According to the protein fractions present as well as from the dynamics of the protein excretion, orthostatism in these five subjects probably originated from organ structural lesions. In the other 14 subjects functional proteinuria was present, but there were differences among them in the dynamics of the appearance and disappearance of the orthostatism, which has been used for arbitrary division of the orthostatic and the so-called march-proteinuria.     

Environmentally related diseases of the urinary tract

Nephrotoxicity from exposure to therapeutic agents and chemicals in the environment and workplace results in a broad spectrum of clinical renal disease that may mimic disorders from other causes. Nephrotoxic agents may, in fact, be responsible for some fraction of renal disease of undetermined etiology. Specific diagnosis and treatment by removal from exposure to the toxic agent is more likely in the early phase of the disorder. Measurement and characterization of proteinuria provides the most sensitive and reliable method of early detection. Increased urinary excretion of serum proteins with molecular weight in excess of 50,000, such as albumin and transferrin, is an early indicator of glomerular injury. Low-molecular-weight proteinuria (beta 2-microglobulin or retinol-binding protein) and enzymuria, particularly excretion of NAG, are sensitive indicators of renal tubular cell injury. Tests that reflect hypersensitivity reactions are often indicative of immunologically mediated nephrotoxicity but are not specific for the kidney. Cancers of the kidney and urinary bladder appear to be increasing and are most common among the socially active and affluent. Susceptibility of the urinary tract to toxicity and carcinogenicity reflect contact of excreted toxins with the epithelial cells of nephrons and urinary bladder.     

Differentiation of glomerular, tubular, and normal proteinuria: determinations of urinary excretion of β2-microglobulin, albumin, and total protein

A low molecular weight beta(2)-globulin (beta(2)-microglobulin), albumin, and total protein were measured in concentrated 24-hr urine specimens from 20 healthy subjects and 30 patients with clinical proteinuria of glomerular or tubular type. Classification of proteinuria was made on the basis of clinical diagnosis and size distribution of urinary proteins after gel chromatography. The molecular radii (Stokes' radii) of beta(2)-microglobulin and albumin, estimated by gel chromatography, were 15 A and 35 A.The average 24-hr urinary excretion in healthy subjects was 0.12 mg for beta(2)-microglobulin, 10 mg for albumin, and 80 mg for total protein. The patients with renal glomerular disorders had normal or only somewhat increased excretion of beta(2)-microglobulin, despite considerably increased excretion of albumin and total protein. Most of the patients with tubular dysfunction excreted large amounts of beta(2)-microglobulin, although they excreted normal or only slightly increased amounts of albumin and only moderately increased quantities of total protein. Consequently, the ratio or urinary albumin/urinary beta(2)-microglobulin was high in glomerular proteinuria (1100: 14,200), intermediate in normal proteinuria (33: 163), and low in tubular proteinuria (1.0: 13.3). Determinations of urinary clearances of beta(2)-microglobulin and albumin in four healthy subjects and 11 patients indicated that increased excretions of the two proteins were associated with increased clearances. The results suggest that quantitative determinations of urinary beta(2)-microglobulin and urinary albumin may be useful for detecting disorders of the renal handling of plasma proteins. The findings also seem to suggest a selective tubular reabsorption of the two proteins.Estimates on sera revealed a close correlation between serum levels of beta(2)-microglobulin and creatinine and also a greatly raised serum concentration of beta(2)-microglobulin after bilateral nephrectomy.     

Analytical characterization of the urinary proteins from sixty patients with monoclonal gammopathies

The urinary proteins from 60 patients with monoclonal gammopathies were characterized by the combination of conventional cellulose acetate electrophoresis, sodium dodecyl sulphate—polyacrylamide gel electrophoresis, and immunoelectrophoresis. Bence—Jones proteinuria was found in 60% of the patients. Non-specific proteinuria was found in 23 of the 37 patients with Bence—Jones proteinuria, and in 9 patients who did not eliminate monoclonal free light chains in their urines. In most patients this non-specific proteinuria followed a pattern suggestive of predominantly glomerular compromise. κ-chains were eliminated predominantly in the monomeric form, while λ-chains were found to exist mainly as dimers. In two patients the monomer:dimer ratio changed during observation perhaps as a manifestation of “escape” of the neoplastic clone from treatment.     

A practicable two-dimensional electrophoretic method for routine analysis of urinary proteins

A two dimensional electrophoretic method is described for the routine clinical analysis of urinary proteins. Cellulose acetate electrophoresis is used for the first dimension, and SDS (sodium dodecyl sulphate) electrophoresis for the second dimension, the latter being performed together with gel staining (Coomassie Blue) on the "Phast System". The separation media are supplied as "ready-to-use" materials. The method is reliable and reproducible, and is complete within 100 minutes. The resulting two-dimensional pattern of major proteinuria constituents is evaluated visually from the distribution according to molecular weight (second dimension) and from the five zone pattern of cellulose acetate electrophoresis (first dimension). Certain "marker" proteins specific for certain pathological changes, as well as certain characteristic changes in protein spot constellation, can be more easily recognized and evaluated than in one-dimensional SDS electrophoresis.     

Effects of anti-proteinuric therapy with angiotensin-converting-enzyme inhibition on renal protein catabolism in the adriamycin-induced nephrotic rat

A direct consequence of glomerular protein leakage is an increased exposure of proximal tubular cells to proteins. The aim of the present study was to examine whether chronic proteinuria affects the tubular handling of proteins and whether anti-proteinuric therapy by angiotensin-converting-enzyme (ACE) inhibition restores this tubular function. Renal uptake and catabolic rate of the low-molecular-weight protein (LMWP) myoglobin was determined in anaesthetized control and adriamycin-induced nephrotic rats by external counting after radiolabelling. Proteinuria correlated with the uptake as well as the catabolism of myoglobin. The higher the proteinuria, the lower was the renal uptake of myoglobin (r =0.72, P =0.002). Also, the catabolic rate of myoglobin (r =0.80, P =0.0002) was lower with increasing severity of proteinuria. During treatment with the ACE inhibitor lisinopril, proteinuria was lowered by 79+/-9% (mean+/-S.E.M.). Renal uptake and catabolic rate of the LMWP were not restored by ACE inhibition. The catabolic rate of myoglobin was even decreased further with 48+/-5% compared with pretreatment levels. In summary, adriamycin-induced proteinuria is associated with a lower uptake and a lower catabolic rate of LMWP in the proximal tubule. ACE inhibition lowers proteinuria, but does not restore the affected LMWP uptake and rate of catabolism. The rate of LMWP catabolism is even decreased further. In contrast, the urinary excretion of N -acetyl glucosaminidase, the tubular marker of toxicity, was effectively returned to normal levels during ACE inhibition. Taken together, the data suggest that proteinuria is toxic for the proximal tubular cells and that ACE inhibition protects the remaining functional tubular cells directly against destruction through decreasing hypercatabolism.