Validation of 5 routine assays for serum free testosterone with a candidate reference measurement procedure based on ultrafiltration and isotope dilution-gas chromatography-mass spectrometry

BACKGROUND/METHOD: The analytical validity of free testosterone (FTe) analog immunoassays is subject to much controversy. We revisited the validation of 4 analog assays and 1 FTe calculation procedure with a metrologically traceable reference measurement procedure (RMP) based on ultrafiltration and isotope dilution-mass spectrometry for direct measurement of Te in the ultrafiltrate. To this end, we performed split-sample measurements of 40 male sera. RESULTS: Deming regression showed that 3 of the immunoassays had moderate to good correlation (0.8474 < or = r < or = 0.9241) with the RMP; however, the slope was markedly below 1. The FTe calculation procedure was in good agreement with this result. The Sy/x values for all assays were higher than the combined imprecision values, which indicate their susceptibility to matrix-related effects. CONCLUSIONS: The study demonstrated substantial differences in analytical quality of FTe assays; however, the results suggested that after extending the validation with a larger variety of samples, recalibration of some analog assays might be worth further investigation.     

Polyanions and Glucose as Osmotic Agents in Simulated Peritoneal Dialysis

Peritoneal dialysis solutions contain glucose as an osmotic agent to obtain ultrafiltration. Due to rapid absorption, glucose does not sustain high ultrafiltration during long exchanges. Nonabsorbable polyanions might be effective as osmotic agents when suspended in electrolyte solution. Concentrations of freely diffusible ions should be in Gibbs-Donnan equilibrium with plasma electrolytes. The ideal proportion of diffusable to polymerbound cation concentrations is unknown. To obtain concentrations of free ions in equilibrium with plasma, it is assumed that the polymer solution dialyzed against a polyelectrolyte solution of the desired composition (with hydraulic pressure higher on the polymer side) will approach the same thermodynamic activity as the electrolyte solution. Subsequently, if transmembrane pressure is released, osmotic ultrafiltration will occur in proportion to the hydrostatic pressure applied during polymer solution preparation. Polyacrylate solution so prepared was compared with a commercial dextrose dialysis solution in an in vitro simulation of peritoneal dialysis. With dwell times up to 24 h, sustained ultrafiltration with polymer was observed, whereas, with dextrose, ultrafiltration ceased after 8 h. Concentrations of diffusible bivalent cations in polyacrylate were lower than intended due to avid polymer complexing; however, dextran sulfate solutions were developed to contain desired concentrations of diffusible electrolytes. The conclusion is that some polymer solutions might be useful in clinical settings when high sustained ultrafiltration is needed.     

Ultrafiltration and Dry Weight—What Are the Cardiovascular Effects?

Long-term prognosis in dialysis is poor compared to that in healthy control persons. A worsening of the prognosis is noted especially for patients who at initiation of dialysis have congestive heart failure, ischemic heart disease, or left ventricular dysfunction or hypertrophy. This is the main reason that cardiovascular causes are the most common for morbidity in these patients. The weight obtained when normal urine output is present is the dry weight. With reduced ability to excrete the volume by the kidneys in end-stage renal disease (ESRD), the body will retain water and the patient will gain weight. This extra weight is due to volume overload. While volume overload may induce a rise in blood pressure, if the heart is in acceptable condition, a fast removal of fluid by ultrafiltration (UF) during dialysis may instead cause hypotension. Ultrafiltration failure in peritoneal dialysis (PD) patients may lead to successive water retention and overhydration with subsequent cardiac failure, while volume overload may occur over a few days in hemodialysis (HD) patients. Anemia or even too-high hematocrit may impair cardiac function further and worsen conditions caused by wrong dry weight. Thus, during long-term and sustained volume overload, left ventricular (LV) hypertrophy will occur in an eccentric manner. A sustained overload then may lead to cell death and LV dilatation and, eventually, systolic dysfunction. Once a severe left ventricular dilatation has developed, the blood pressure may decrease during volume overload. A worsened prognosis is seen if malnutrition and low albumin levels are present. Volume overload necessitates ultrafiltration to achieve dry weight. Thereby, volume contraction contributes to exaggerated stimulation of or response to activation of the RAS and alpha-adrenergic sympathetic systems. If ultrafiltration goes beyond these compensatory mechanisms, hypotension will occur and increase the risk for hypoperfusion of vital organs. Such episodes may cause cardiac morbidity, aspiration pneumonia, vascular access closure, or neurological complications (seizures, cerebral infarction), besides a more rapid lowering of residual renal function. Preventive measures are, first, finding the right dry weight; second, minimizing interdialytic weight gain; third, optimizing the target for hemoglobin (110-120 g/l); fourth, lowering dialysate calcium (1.25 mmol/l); and fifth, eventually using higher dialysate potassium if long dialyses are performed.     

Non-invasive intravascular monitoring in the pediatric hemodialysis population

Assessment of dry weight in pediatric hemodialysis (HD) patients is difficult, since small fluid shifts may result in dialysis-associated morbidity (DAM) and children may not verbalize complaints. Achieving dry weight is critical since chronic fluid overload can result in hypertension and left ventricular hypertrophy. To determine if non-invasive monitoring of hematocrit (NIVM) is useful in preventing DAM in pediatric HD patients, we reviewed 200 HD treatments performed with or without NIVM (no NIVM). DAM was defined as an ”event” (e.g., hypotension, headache, cramping) that required nursing intervention. Patient age, weight, and gender were similar in both groups. Desired ultrafiltration was obtained in both groups. The event rate was lower in NIVM than no NIVM for all treatments (0.22 vs. 0.3, P=0.07) and significantly lower in patients <35 kg (0.25 vs. 0.47, P=0.01). The second event rate (fraction of treatments with one event that had a subsequent event occurring at least 15 min later) was lower with NIVM (P<0.01). For the NIVM group, events in the first 90 min occurred when blood volume changed >8% per hour; 71% of events (43/60) at 90–240 min occurred when blood volume changed >4% per hour. NIVM decreases DAM in pediatric HD patients, especially those <35 kg. Ultrafiltration with blood volume change <8% per hour is safe in the 1st h and <4% after 1 h reduces DAM in children. Received: 5 June 2000 / Revised: 29 August 2000 / Accepted: 6 September 2000     

Inter-laboratory comparison of DNA preservation in archival paraffin-embedded human brain tissue from participating centres on four continents

DNA extracted from formalin-fixed and paraffin-embedded brain tissue is known to contain as yet ill-characterized inhibitors of the PCR process. As part of a project that aims to clarify the role of mitochondrial DNA sequence variation in human neurodegenerative diseases using DNA from various ethnic backgrounds, we have investigated factors that influence the preservation of archival DNA and its suitability for PCR. In this study, neuropathological tissue samples were analysed that had been routinely processed in 18 international centres on four continents. Following DNA extraction, PCR amplification of mitochondrial and nuclear DNA sequences was performed with and without additional purification of the template DNA. In addition, the DNA used for PCR was analysed by HPLC. Phosphate-buffered formalin proved to be a superior fixative compared with unbuffered aldehyde: DNA extraction resulted in greater yields, the molecular weight of the isolated DNA was higher and PCR was more successful. PCR inhibitors were identified as (1) high concentrations of small (<300 bp) DNA fragments that competitively compete with template DNA and (2) contaminants of the DNA template solution including denatured protein that cannot be completely removed by phenolic extraction. HPLC analysis did not reveal significant qualitative differences between DNA isolated from fresh-frozen tissue samples and DNA recovered from formalin-fixed, paraffin-embedded brain tissue. The fact that DNA could be amplified from the majority of tissue specimens in this study suggests that rare diseases and diseases where ethnic background plays an important role can be sampled for genetic polymorphism analysis on a global scale using archival neuropathological collections. Electronic Publication     

复合超滤对小儿先天性心脏病术后肺功能的影响

目的探讨复合应用改良超滤和零平衡超滤对改善小儿先天性心脏病体外循环(extracorporeal circulation,ECC)术后肺功能的临床效果。方法60例行室间隔缺损修补术(VSD)的患儿随机分为四组:常规超滤组(CUF组,n=15)、改良超滤组(MUF组,n=15)、零平衡超滤组(ZUF组,n=15)和改良超滤 零平衡超滤组(M Z组,n=15)。分别检测各组围术期红细胞压积(Hct),呼吸功能氧合指数(OI),肺泡-动脉血氧分压差(P(A-a)O2),呼吸机辅助时间(MAT)及炎性介质肿瘤坏死因子α(TNF-α)和白细胞介素-6(IL-6)的浓度。结果术后各组Hct无显著性差异(P>0.05);M Z组、ZUF组和MUF组术后1 h,6 h,12 h,24 h OI高于CUF组(P<0.05),而M Z组术后1 h,6 h,12 h,24 h P(A-a)O2较ZUF组、MUF组和CUF组低(P<0.05);M Z组术后呼吸机支持时间较MUF组、ZUF组和CUF组短(P<0.05);M Z组和ZUF组停机及术后2 h,12 h,24 h TNF-α和IL-6浓度较MUF组和CUF组明显降低(P<0.05)。结论零平衡超滤加改良超滤较单一超滤方法能较好地改善小儿先天性心脏病患者术后肺功能,降低体内炎性介质浓度。     

Technical Requirements for Rapid High-Efficiency Therapies

Significant reductions in treatment time are possible with rapid high-efficiency hemodialysis and hemodiafiltration, provided the therapies are implemented so as to maintain adequacy of solute and fluid removal without compromising patient comfort. The key technical elements necessary for such implementation include high blood flow rates, higher efficiency dialyzers/diafilters, ultrafiltration control systems, and bicarbonate as the buffer source. In addition, hemodiafiltration requires schemes to ensure sterility and nonpyrogenicity of the infusion fluid and appropriate balancing of the rates of ultrafiltration and reinfusion. In general, rapid high-efficiency therapies are technically more complex than standard therapy, and rapid hemodiafiltration appears to be more complex than rapid hemodialysis. The technology required for rapid hemodialysis currently exists, but it needs to be fine-tuned and integrated for routine application.     

血液透析中发生血压异常的情况分析

目的 观察不同年龄组血液透析中高血压及低血压的发生率，对有关因素及与超滤量(UFV)和每公斤体重超滤量(UFV／W)的关系进行分析。方法 对我院常规透析的加例患者的1575例次的透析进行观察分析。结果 老年组患者高血压发生率小于非老年组，而低血压发生率较非老年组显著提高。糖尿病、冠心病、心包积液、充血性心力衰竭等并发症的发生率也大于非老年组。老年组透析期间发生低血压时较其正常血压时UFV／W明显增高，UFV无显著性差异。非老年组透析期间发生低血压时较其正常血压时UFV明显增高，UFV／W无显著差异。两组血压增高时分别与其正常血压时UFV及IFV／W无明显差异。结论 血液透析中UFV或UFV／W过高是血压降低的关键，尤其是老年患者。     

腹膜转运特性明显影响艾考糊精腹透液的超滤量

目的 观察不同腹膜转运特性的患者使用7.5%艾考糊精腹透液长时间留腹后的超滤量。 方法 采用前瞻性、多中心、随机、双盲和平行对照临床研究的亚组分析。连续非卧床腹膜透析（CAPD）患者根据腹透液肌酐与血肌酐比值（D/Pcr）和Twardoski的评判标准,分为高转运、高平均转运、低平均转运和低转运4组。患者分别使用7.5%艾考糊精透析液或2.5%葡萄糖 Dianeal?誖 PD-2、PD-4透析液,疗程4周。比较各组超滤量。 结果 共201例CAPD患者入选,其中艾考糊精组98例,葡萄糖组103例;男96例,女105例;年龄（56.1±13.7）岁（18～81岁）。198例进行了腹膜平衡试验,其中高转运患者24例（12.1%）,高平均转运72例（36.2%）,低平均转运81例（40.7%）,低转运21例（11.0%）。治疗第4周,高转运、高平均转运和低平均转运患者中,艾考糊精组超滤量显著升高,无论与基线值还是与葡萄糖组比较,差异都有统计学意义;而低转运患者中,艾考糊精组超滤量高于葡萄糖组,但差异无统计学意义。相关分析显示,艾考糊精腹透液产生的超滤量与D/Pcr呈正相关（R2 = 0.1681,P < 0.01）,而葡萄糖透析液产生的超滤量与D/Pcr呈负相关（R2 = 0.0949,P < 0.01）。 结论 7.5%艾考糊精腹透液可显著改善CAPD患者的超滤量和腹膜肌酐清除率,尤其在腹膜高转运、高平均转运和低平均转运患者,其作用显著优于葡萄糖腹透液。     

超滤法纯化三七总皂苷的工艺研究

目的探究超滤分离技术纯化三七总皂苷.方法三七经大孔树脂纯化,氧化铝、活性碳精制,在浓缩干燥前增加超滤工艺,并对超滤液的可见异物、有效成分含量、截流率、膜通量测定.结果选用卷式截留相对分子量100KD的聚醚砜膜作为中试超滤膜较适合.结论所选择的膜分离技术纯化三七总皂苷,可减少超滤液中的可见异物数量,提高澄明度.