国产放射免疫试剂与Abbott放射免疫试剂检测乙型肝炎病毒感染指征的比较

本文报告了国产乙型肝炎病毒(HBV)RIA和Abbott RIA诊断试剂盒检测质量的比较研究。表明两试剂滴定HBsAg、抗-HBs、抗-HBc阳性血清的敏感性相似,抗-HBs mIU/ml定量检测结果相符。证明国产RIA试剂质量是可靠的。但是,国产试剂S/N值曲线的陡度和标本的阳性检出率较Abbott试剂为高,可能与试剂出厂后周转期短有关。适当提高国产试剂阳性判定标准后,与Abbott试剂的检测符合率可达90％以上。     

中和试验和ELISA法检测育龄妇女血清麻疹抗体的比较

目的:研究中和试验和ELISA两种麻疹抗体检测方法结果的相关性,并初步探讨新生儿对麻疹易感的原因.方法:采集本院住院的85例小于9月龄麻疹患儿母亲血清,同时采集13位幼年患过麻疹的育龄妇女血清作为对照.通过细胞培养测定血清中和抗体滴度,同时使用ELISA诊断试剂定量检测麻疹IgG抗体,统计学分析比较两种检测结果的相关性.结果:98例血清ELISA法和中和试验法两者均阳性89例,总符合率为90.82%,两者结果不符合主要见于抗体浓度较低的标本;两种检测方法麻疹抗体检测结果具有正相关性(R=0.8285,P＜0.001).当中和抗体滴度大于1:16时,ELISA抗体浓度均高于200 mIU/ml;而当ELISA抗体浓度均高于700 mIU/ml,中和抗体均阳性.患儿患病时,其母亲麻疹ELISA抗体平均水平633 mIU/ml,平均中和抗体滴度1:37,而幼年有麻疹史的同龄妇女麻疹ELISA抗体平均水平为1498 mIU/ml,中和抗体滴度为1:182;两组差异有统计学意义(P＜0.05).结论:中和试验和ELISA麻疹抗体检测结果具有良好的相关性;ELISA麻疹抗体浓度高于700mIU/ml是麻疹中和抗体阳性的可靠指标.婴幼儿麻疹易感性增加与其母亲麻疹抗体低下有关.     

乙型肝炎病毒基因重组疫苗免疫儿童成人效果及阻断母婴传播的研究

以中试连续生产的6批乙肝基因重组疫苗(R—Hepavac—B)免疫儿童和成人,每剂10μg,3批苗按0、1、6月,另3批苗按0、1、2月方案接种。每批分别接种8～10岁儿童51～66人,6批共363人,18～20岁成人46～52人,6批共287人。同时用两批血源苗作对照,分别接种儿童共85人,成人84人。又用其中89—10批HBsHg含量为20μg/ml的疫苗,按20μg×3,0、1、6月方案接种HBsAg和HBeAg双阳性母亲所生婴儿36名,HBsAg阳性母亲的婴儿19名。结果显示,10μg重组疫苗3针后,无论儿童和成人其抗HBs抗体阳转率均为100％,而对照,儿童成人各有1例未阳转。儿童按0、1、6方案接种者,其抗体GMT为363.8～470.5mIU,按0、1、2方案接种者,为150.8～195.5mIU。前者高于血源苗对照,而后者持平。成人0、1、6方案GMT为189.4～247.2mIU,0、1、2方案为87.9～96.3mIU,均略低于血源苗对照.经复测,此批血源苗含量为15μg/ml,高于基因苗含量,可能是造成以上结果的原因。母婴阻断结果显示,双阳性母亲子女36人,3针免后86.1％(32/36)的婴儿获得保护。单阳性母亲子女19人全部获得保护。     

母源性抗-HBs对新生儿接种乙型肝炎疫苗的远期影响

目的:观察新生儿体内母源性乙型肝炎表面抗体(抗-HBs)对乙型肝炎疫苗接种后抗体应答的远期影响。方法:2006年10月-2007年1月在南京大学医学院附属鼓楼医院产前检查并住院分娩的单胎足月妊娠妇女中,选择抗-HBs阳性孕妇32例,抗-HBs阴性孕妇32例,其新生儿均按0、1、6方案接种重组(酵母)乙型肝炎疫苗,检测两组儿童乙型肝炎疫苗第3针接种后2年血清抗-HBs浓度。结果:新生儿接种乙型肝炎疫苗第3针后2年,母源性抗-HBs阴性组与阳性组儿童抗-HBs阳性率分别为90.6%与87.5%,其抗-HBs几何平均浓度分别为73.48mIU/ml与75.49mIU/ml,两组间抗-HBs阳性率与GMC的差异均无统计学意义(P=1.000,P=0.778);6例母源性抗-HBs>1000mIU/ml儿童中1例抗-HBs转阴(16.7%),而母源性抗-HBs<1000mIU/ml组和母源性抗-HBs组的儿童抗-HBs转阴率分别仅为7.69%和11.1%,但差异无统计学意(P=0.811)。结论:新生儿目前按照0,1,6方案接种乙型肝炎疫苗,能够有效保护其抵抗乙型肝炎病毒的感染;母源性抗-HBs对新生儿接种乙型肝炎疫苗...     

HBsAg阴性母亲的新生儿乙型肝炎疫苗免疫后抗-HBs的动态研究

HBsAg阴性母亲的新生儿,按0、1、6个月程序接种3剂10μg乙型肝炎血源疫苗,在第一针免疫后9～48个月观察抗-HBs阳转率,发现免后12个月阳转率最高达94.23％;至48个月为81.62％。48个月后抗-HBs GMT仍有263.5mIU/ml,提示此期间不必加强免疫。抗-HBs滴度(mIU/ml)的动态规律是由高滴度(>1000mIU/ml)向低滴度推移。     

池养雄性白Ji豚性自慰行为与血清睾酮浓度之间的关系

自1996年3月至1998年12月,作者采用放射免疫分析法每月1次测定雄性白Ji豚（Lipotes vexillifer)“淇淇”血清睾酮浓度。春季每天1次,其它季节每三天1次,每次1h,用目标动物取样法观察“淇淇”的行为。结果发现,性自慰（腹面贴池壁、伸出阴茎）行为的发生频率和血清睾酮激素水平都有周期性变化,春季为高峰期,秋季有1个小峰,性自慰行为与血清睾酮激素浓度两者的周期较为一致。从性自慰行为和血清睾酮激素水平季节变化,推测出“淇淇”性腺活动的周期性变化,活动期为2－10月,血清睾酮浓度为2．48－9．66ng/ml。春季高峰在4月前后,血清睾酮浓度为3．44－9．66ng/ml;秋季小峰在10月,血清睾酮浓度为3．33－4．25ng/ml;11月至次年1月为性腺活动相对静止期,血清睾酮浓度为1．24－2．85ng/ml。白Ji豚是一种季节性繁殖动物,繁殖高峰期在4月前后。     

树状多节孢发酵生产紫杉醇工艺条件的初步研究

研究了树状多节孢HQD33内生真菌融合子TPF－1瓶发酵工艺条件,进行了2．8L和10L通用式机械搅拌罐的发酵试验。结果表明,HQD33适宜发酵工艺条件是：发酵时间16－18d,培养基中蔗糖、苯丙氨酸、醋酸钠、酪氨酸,2,4-氯苯氧乙酸和亚油酸的加量分别为180g/L、1mg/L,1.5g/L,15mg/L,5mg/L和15mg/L,摇瓶装置为150ml/500ml,在此条件下摇瓶发酵液中紫杉醇平均含量为448．52μg/L;2.8L和10L罐发酵液中紫杉醇含量达406．95和395．12μg/L（平均值）。     

bcl-2癌基因蛋白在甲状腺癌中的表达及其意义

利用免疫组织化学技术,对６４例甲状腺癌进行了ｂｃｌ－２蛋白表达的检测,同时进行ｐ５３蛋白的对照检测。结果显示,甲状腺癌中ｂｃｌ－２蛋白阳性表达率为８１．３％（５２／６４）,但未分化癌无阳性表达。ｐ５３蛋白在甲状腺癌的阳性表达率为２０．３％（１３／６４）,而未分化癌全部为阳性表达。两种抗体在甲状腺癌的阳性表达率有显著性差异（Ｐ＜０．０１）。结果提示ｂｃｌ－２蛋白在甲状腺癌的表达与肿瘤细胞的分化程度有关,并与ｐ５３蛋白呈反比关系,ｂｃｌ－２与ｐ５３蛋白表达的不同分布可作为判断甲状腺癌预后的一个重要参考指标     

大鼠胰岛分离条件的优化

目的:优化大鼠胰岛分离纯化的条件,为胰岛移植实验奠定基础。方法:通过胆总管灌注胶原酶P来消化大鼠胰腺,分离胰岛,采用不连续密度梯度Ficoll离心法纯化胰岛,观察胶原酶浓度、消化时间以及大鼠体重对胰岛分离结果的影响。双硫腙染色鉴定胰岛,丫啶橙/碘丙啶染色鉴定胰岛细胞活率,糖刺激胰岛素释放试验评价胰岛功能。结果:胶原酶浓度、消化时间以及大鼠体重对胰岛分离结果有重要影响。1mg/ml胶原酶P在37℃静止消化45分钟条件下,胰岛分离效果最佳,效果较其他酶浓度和消化时间条件下好(P＜0．05)。体重350g的大鼠的胰岛收获量778．33±80．21IEQ/胰腺,而体重250g的大鼠的胰岛收获量655．00±56．56 IEQ/胰腺(P＜0．05)。优化条件下分离的胰岛其纯度＞90%,胰岛细胞活率＞90%,低糖(2．8mmol/L)、高糖(16．7mmol/L)刺激胰岛素释放分别为(5．40±1．75)mIU/L/30IEQ,(12．27±2．55)mIU/L/30IEQ(P＜0．05),刺激指数为2．33±0．29。结论:胶原酶浓度、消化时间以及大鼠体重影响胰岛分离结果,优化分离条件可改善大鼠胰岛分离结果。     

白术提取物对六价铬致人外周血淋巴细胞 抗突变作用研究

目的：探讨白术提取物抗六价铬的致突变作用。方法：采用人外周血淋巴细胞程序外DNA合成（UDS）试验检测了0．03g、0．015g和0．0075／ml白术提取物对重铬酸钾（含Cr^6＋0．31ug/ml）所致淋巴细胞程序外DNA合成的影响。同时检测白术提取物与重铬酸钾的加入顺序对UDS的影响。结果：阳性对照（重铬酸钾,含CCr^6＋0.31ug/ml）组CPM值显著高于阴性对照（蒸馏水）组,P〈0．01。白术提取物与重铬酸钾同时加入以及两者间隔2h先后加入均显示白术提取物浓度与CPM值呈显著负相关（P〈0.05）,在上述不同加入方法中,白术提取物的各浓度均显示CPM值显著低于阳性对照。结论：白术提取物具有抑制Cr^6＋的致突变作用。     

Evaluation of autoantibodies against oxidized LDL and antioxidant status in top soccer and basketball players after 4 months of competition

Antioxidant status and titers of autoantibodies against oxidized low-density lipoproteins (ox-LDL-Ab) were investigated in top soccer (S; n = 21, age 24.6 +/- 4.3 years) and basketball (B; n 3,000 mIU/ml) in ox-LDL-Ab were found in half the players (12S and 4B) with a maximum reaching 6000 mIU/ml (normal range: 200-600 mIU/ml), showing an in vivo LDL oxidation. There was no correlation between ox-LDL-Ab titers and chlolesterol, LDL cholesterol, or antioxidant levels. Nevertheless, plasma vitamin C concentration was lower in athletes having high levels of ox-LDL-Ab when compared with those with normal levels (8.49 +/- 3.14 mirogram/ml vs. 10.39 +/- 2.55 microgram/ml), but this difference was not statistically significant. In conclusion, our data suggest that potential atherogenic and cardiovascular risks as reflected by high titers in ox-LDL-Ab may exist in some top athletes despite a nonaltered antioxidant status.     

Radioimmunoassay of nafarelin ([ 6-(3-(2-naphthyl)-D-alanine)]-luteinizing hormone-releasing hormone) in plasma or serum

A procedure which is suitable for the radioimmunoassay (RIA) of nafarelin [( 6-(3-(2-naphthyl)-D-alanine)]-luteinizing hormone-releasing hormone) in plasma or serum at concentrations as low as 50 pg/ml is described. Antiserum was prepared by replacing the pyroglutamyl portion of nafarelin with glutaric acid, coupling the product to keyhole limpet hemocyanin, and immunizing rabbits with the resulting conjugate. At a dilution of 1:30,000 the binding was approximately 50%. The antibodies did not cross react with luteinizing hormone-releasing hormone. For RIA, 125I-labeled analyte was used as the tracer and charcoal was used to separate the free and the bound fractions. No purification of samples was required prior to RIA. Accuracy of the method was assessed by adding known quantities of nafarelin to nafarelin-free plasma and determining the ratio of measured to added analyte. Linear regression analysis for the concentration range 0.050-5.00 ng/ml yielded a regression equation of y = 1.01x - 0.066 and a correlation coefficient of 0.997. At 0.050 ng/ml the CV was 11.3% (interassay). Additional validation was obtained from an in vivo study in which [3H]nafarelin was administered to monkeys and plasma profiles were determined by RIA, by high-performance liquid chromatography (HPLC), and by an HPLC-radiochemical method. The results obtained by RIA agreed well with those obtained by the HPLC methods.     

Critical considerations in the development of an assay for sulfidopeptide leukotrienes in plasma

A sensitive and specific assay has been developed for measurement of total sulfidopeptide leukotrienes (LT) in plasma. LTC4 and LTD4 in plasma are converted to LTE4 which is then extracted by C18 Sep-Pak binding and elution. Total LTE4 is resolved by reverse phase high performance liquid chromatography (RP-HPLC) and quantitated by radioimmunoassay (RIA). A [3H]LTE4 internal standard is added to the starting plasma sample to allow overall recovery to be calculated and to define the fractions from RP-HPLC to be assayed for LTE4-like immunoreactivity. The correlation between the measured increase in LTE4 concentration after addition of incremental amounts of LTC4 and LTE4 to plasma was 0.989 and 0.978, respectively, with slopes of 1.05 and 1.11. Addition of 51 pg/ml LTE4 to 5 ml plasma was detectable; the measured increase was 48 +/- 12 pg/ml (mean +/- SE, n = 7). The intra-assay coefficient of variation for 341 pg/ml of added LTC4 was 3.2% (n = 6). Sulfidopeptide leukotrienes could not be detected in blood samples taken from 12 normal volunteers in whom the theoretical detection limit, calculated from the sensitivity of the RIA, the overall recovery of LTE4, and the volume of plasma extracted, was 83 +/- 4 pg LTE4/ml plasma (0.19 +/- 0.01 pmol sulfidopeptide leukotriene/ml plasma; mean +/- SE).     

Effect of a melatonin implant on the circadian variation of plasma prolactin and rectal temperature in newborn sheep.

Plasma prolactin and rectal temperature show a circadian rhythm in newborn sheep raised under continuous light. Melatonin lowers the concentration of plasma prolactin but it is not known if it affects its circadian rhythm. To detect whether melatonin acts on the circadian system we studied the effect of a subcutaneous melatonin implant in the circadian rhythms of prolactin and rectal temperature in newborn lambs raised under continuous light. We placed catheters in the pedal artery and vein in 9 newborn lambs (2-5 days of age). A subcutaneous melatonin implant was placed in 4 of the lambs at 9-12 days of age. Blood samples and rectal temperature measurements were obtained hourly for a period of 24 h, 11-15 days after the implant, at 20-27 days of age. To avoid interferences of heparin in our melatonin assay, serum melatonin concentration was measured before and during the implant in three additional newborns. Prolactin and melatonin were measured by RIA. Melatonin concentrations were 52.8 +/- 45.9 pg/ml (day) and 315.5 +/- 77.0 pg/ml (night) before treatment (SEM, P less than 0.001), and increased to 594.1 +/- 54.5 pg/ml after placing the implant (there was no difference in melatonin concentration between day and night during the time that the implant was in place). Melatonin had no effect on rectal temperature or its rhythm, but decreased basal plasma prolactin concentration (control: 97.5 +/- 11.3 ng/ml; treated: 25.1 +/- 2.4 ng/ml, P less than 0.001) and abolished the prolactin circadian rhythm, (Cosinor analysis): control: log prolactin (ng/ml) = 1.8 + 0.26 cos 15 (t - 11.16), p = 0.05; treated: log prolactin (ng/ml) = 1.2 + 0.14 cos 15 (t - 9.43), P = 0.36.     

Identifying Prediabetes Using Fasting Insulin Levels

ObjectiveTo determine whether patients with prediabetes can be accurately and easily identified in clinical settings using a predictive clinical and laboratory model.MethodsThis retrospective study examined demographic and laboratory data from patients who had undergone 2-hour glucose testing for suspected prediabetes or diabetes between 2000 and 2004. Patients who met the diagnostic criteria for diabetes mellitus were excluded. Prediabetes was defined as a fasting glucose concentration ≥ 100 mg/dL and ≤ 125 mg/dL or a 2-hour postprandial glucose concentration ≥ 140 mg/dL and < 200 mg/dL. Multivariate logistic regression was conducted to identify calculated or measured clinical and laboratory attributes that predict the presence of prediabetes, including fasting insulin quartiles, homeostasis model assessment of insulin resistance (HOMA-IR), and quantitative insulin sensitivity check index.ResultsOf 965 patients, 287 (29.7%) had prediabetes. The study population primarily consisted of white, obese, female patients. A multivariate model revealed that compared with the referent lowest quartile of fasting insulin (m = 4.9 [± SD] ± 1.2 mIU/mL), subsequent insulin quartiles increased the likelihood of identifying prediabetes (quartile 2: m = 8.0 ± 0.8 mIU/mL, odds ratio [OR] = 2.076, confidence interval [CI] = 1.241-3.273; quartile 3: m = 12.2 ± 1.7 mIU/mL, OR = 3.151, CI = 1.981-5.015; quartile 4: m = 25.9 ± 12.4 mIU/mL, OR = 5.035, CI = 3.122-8.122). Older age and increased diastolic blood pressure also contributed modestly to this model. Further analysis using the area under the curve revealed that at a fasting insulin level > 9.0 mIU/mL, prediabetes would be correctly identified in 80% of affected patients. A second model revealed that increased HOMA-IR index (OR = 1.303, CI = 1.205-1.410) and older age (OR = 1.037, CI = 1.024-1.05) predicted prediabetes.ConclusionsThe most robust model, which used fasting insulin levels, may provide the most utility as a clinical tool because the highest quartiles suggest significantly greater likelihood of identifying prediabetes. (Endocr Pract. 2010;16:47-52)     

Concurrent enhancement of monocyte immunoregulatory properties and effector functions by recombinant interferon-gamma

Interferon-gamma is a critical factor in the activation of several mononuclear phagocyte effector and immunoregulatory properties. However, it remains uncertain if IFN-gamma is capable of concurrent activation of both functions in the same cell population. Plastic adherent mononuclear cells (80-98% MN by cytochemical criteria) were cultivated in the absence or presence of recombinant interferon-gamma (rIFN-gamma, 0.1-100 U/ml) for 48 hr. MN surface DR antigen was assessed by flow cytometry (EPICS V) after staining with monoclonal antibodies OKIa1 or L243. Exposure to rIFN-gamma (100 U/ml) increased MN surface DR antigen (mean fluorescence intensity) by 80 +/- 20% (P less than 0.01) and 121 +/- 52% (P less than 0.001), respectively, compared to untreated cells. The increase in DR antigen was maximal at 100 U/ml, dependent on protein and RNA synthesis and blocked by agents that increase cAMP levels. IL-1 activity was determined in the mouse thymocyte assay; rIFN-gamma (100 U/ml) increased IL-1 activity in the supernatants of MN cultured in medium alone from 0.5 +/- 0.2 to 7.8 +/- 4.7 U/ml (P less than 0.05), and lipopolysaccharide-stimulated MN from 20.4 +/- 19.1 to 71.7 +/- 38.9 U/ml (P less than 0.05). Following rIFN-gamma exposure, MN stimulation of the AMLR was increased at 6 days (29,269 +/- 5224 vs 13,252 +/- 4938 cpm, P less than 0.01). Spontaneous cytotoxicity (SC) and antibody-dependent cell cytotoxicity (ADCC) were studied in a 51Cr release microculture assay using the human lymphoblastoid cell line CCRF-CEM as target. SC by MN increased linearly as a function of log[rIFN-gamma] for effector:target (E:T) ratios of 5:1 (r = 0.95, P less than 0.01) and 10:1 (r = 0.99, P less than 0.01). ADCC by MN increased following rIFN-gamma exposure (100 U/ml) at E:T ratios of 5:1 (22 +/- 13 to 31 +/- 4%, P less than 0.025) and 10:1 (31 +/- 4 to 38 +/- 4%, P less than 0.01). Thus, rIFN-gamma not only activates MN effector function, but has concurrent stimulatory effects on multiple MN properties critical to immunoregulation.     

Presence of positive feedback in males with Klinefelter's syndrome

This study examined the effect of 17 beta-estradiol (E2) on basal and luteinizing hormone (LH)-releasing hormone (LHRH)-stimulated gonadotropin secretion in 9 patients with Klinefelter's syndrome. Intramuscular injection of E2 (10 micrograms/kg/day during 5 days) induced a rapid decrease in follicle-stimulating hormone (FSH) and LH levels. The maximum suppression was observed on day 7 (D7) for FSH [median 9.7 mIU/ml (range 4.6-37.8) vs. 21.7 mIU/ml (range 12.2-56.9)] and on D2 for LH [median 13.6 mIU/ml (range 6.8-25.2) vs. 21.2 mIU/ml (range 13-54.7)]. E2 concentrations rose and reached their peak values on D3 [median 723 pmol/l (range 517-1,247.8) vs. 110.1 pmol/l (range 68.6-227.5) on D0]. These changes were followed by a subsequent rise in LH on D4 [36.7 mIU/ml (range 19.4-77.7)]. LH response to LHRH was higher during E2 treatment: median value of absolute peaks: 156.3 mIU/ml (range 56.7-188.6) on D4 vs. 64 mIU/ml (range 38.9-131) on DO. These results demonstrate the presence of a positive feedback in patients with Klinefelter's syndrome.     

Electron factors in the properties of cytochrome P-450 and mechanism of activation of molecular oxygen

A quantum-chemical calculation was carried out for the electronic structures of coordination compounds of general formula: FeP(L1)(L2) (P--porphin; L1 = SHCH3, [SCH3]-, [SC6F4H]-; L2 = CO, NO, O2), modeling the active site of cytochrome P450. It was shown that Coulomb repulsion between the electrons of the sulfur lone pair leads to the transfer of the electronic density from the ligands L1 = [SCH3]- or [SC6F4H]- to the porphyrin of/and to the L2 ligand. This explains the origin of the band at 450 nm in the absorption spectra of the complexes of cytochrome P450 with CO, the absence of such a band in those with O2, and the strong activation of dioxygen by cytochrome P450.     

Protective Effect of HLA-B*5701 and HLA-C -35 Genetic Variants in HIV-Positive Caucasians from Northern Poland

Aim of the Study

Association of two HLA class I variants with HIV-1 pretreatment viremia, CD4+ T cell count at the care-entry and CD4+ T cell nadir.

Methods

414 HIV-positive Caucasians (30% women) aged 19-73 years were genotyped for HLA-C -35 (rs9264942) and HLA-B*5701 variants. HIV-1 viral load, as well as CD4+ T cell count at care-entry and nadir, were compared across alleles, genotypes and haplotypes.

Results

HLA-C -35 C/C genotype was found in 17.6% patients, C/T genotype in 48.1%, and T/T genotype in 34.3% patients. HLA-B*5701 variant was present in 5.8% of studied population. HIV plasma viremia in the group with C allele was significantly lower (p=0.0002) compared to T/T group [mean:4.66 log (SD:1.03) vs. 5.07 (SD:0.85) log HIV-RNA copies/ml, respectively], while CD4+ T cell count at baseline was notably higher among C allele carriers compared to T/T homozygotes [median: 318 (IQR:127-537) cells/μl vs. median: 203 (IQR:55-410) cells/μl, respectively] (p=0.0007). Moreover, CD4+ T cell nadir among patients with C allele [median: 205 (IQR:83.5-390) cells/μl] was significantly higher compared to T/T group [median: 133 (IQR:46-328) cells/μl] (p=0.006). Among cases with HLA-B*5701 allele, significantly lower pretreatment viremia and higher baseline CD4+ T cell count were found (mean: 4.08 [SD: 1.2] vs. mean: 4.84 [SD:0.97] log HIV-RNA copies/ml, p=0.003 and 431 vs. 270 cells/μl, p=0.04, respectively) compared to HLA-B*5701 negative individuals. The lowest viremia (mean: 3.85 log [SD:1.3]) HIV-RNA copies/ml and the highest baseline and nadir CD4+ T cell [median: 476 (IQR:304-682) vs. median: 361 (IQR: 205-574) cells/μl, respectively) were found in individuals with HLA-B*5701(+)/HLA-C –35 C/C haplotype.

Conclusions

HLA-C -35 C and HLA-B*5701 allele exert a favorable effect on the immunological (higher baseline and nadir CD4+ T cell count) and virologic (lower pretreatment HIV viral load) variables. This protective effect is additive for the compound HLA-B*5701(+)/HLA-C -35 C/C haplotype.     

Measurement of DNA methylation using stable isotope dilution and gas chromatography-mass spectrometry

A simple, highly selective, and sensitive method has been developed to quantify methylation of DNA extracted from human peripheral blood mononuclear cells. Assay has been performed at nucleobases level. Cytosine and 5-methylcytosine DNA content has been detected by gas chromatography-mass spectrometry using [2-(13)C]cytosine and [2-(13)C]5-methylcytosine as internal standards. The methylation level has been calculated as 5-methylcytosine/total cytosine ratio. The working range selected on calibration curve, obtained by evaluation of standards and matrix-added standards measurements, is suitable for 5 microg DNA analysis. In this range, healthy human DNA methylation percentage is within 5-6%.