鲎试验和家兔法测定干扰素和白蛋白中内毒素的比较

本文分别用鲎试验和家兔热原法对16批人白细胞干扰素及5批人血浆白蛋白中的内毒素进行了检测。两种方法检测人白细胞干扰素和人血浆白蛋白的符合率分别为81.75%和80%;阴性符合率为100%。     

利用废血浆代替人血清诱生干扰素

分离白细胞时所得废血浆,经处理后可代替人血清生产干扰素。通过16批对比试验,干扰素效价没有明显差别。     

精制人白细胞干扰素口含片的研制

采用空白粒子-喷雾法，制备三批稳定性良好的精制人白细胞干扰素口含片。经中国药品生物制品检定所检定，该制剂外观性状、水分、pH值、崩解时限、干扰素效价及均匀度、细菌学检验等指标均符合《精制人白细胞干扰素日含片制造与检定暂行规程》要求。     

人白细胞干扰素和基因工程干扰素α—1b治疗慢性乙型肝炎

目的：比较人白细胞干扰素和基因工程干扰素α－１ｂ治疗慢性乙型肝炎（ＣＨＢ）疗效和不良反应。方法：ＣＨＢ病人８３例（男性６９例，女性１４例，年龄３２±ｓ８ａ）。Ａ组２９例，用ＬＩＦＮ ３ＭＵ；Ｂ组２７例，用ｒＩＦＮα－１ｂ３ＭＵ，Ｃ组２７例用人血浆白蛋白冻干制品１０ｇ，３组均ｉｍ，ｑｄ×４ｗｋ，继以ｑｏｄ×８ｗｋ。结果：３组ＨＢｅＡｇ阴转率分别为３８％、４１％、７％；ＨＢＶ－ＤＮＡ阴转率分别为３８％     

注射用重组人血清白蛋白-干扰素α2b融合蛋白生物学活性检测方法学研究

目的通过对注射用重组人血清白蛋白-干扰素α2b融合蛋白的生物学活性检测方法进行研究,建立该药物的生物学活性检测方法。方法参考重组人干扰素生物学活性测定法,以人羊膜细胞(WISH)病变抑制法开展注射用重组人血清白蛋白-干扰素α2b融合蛋白的生物学活性检测方法研究。结果该方法的专属性好,精密度试验中生物学活性相对标准偏差(RSD)为10.6%,准确性试验平均回收率98.2%,RSD为9.0%,符合生物制品质量检验方法要求。结论本研究制定的实验方法准确、可靠,适用于注射用重组人血清白蛋白-干扰素α2b融合蛋白的生物学活性检测。     

光阻法检测人血白蛋白、静注人免疫球蛋白中的不溶性微粒

目的对人血白蛋白、静注人免疫球蛋白中的不溶性微粒进行检查研究。方法按照《中国药典》2010年版三部中不溶性微粒检查法-光阻法进行检测。结果样品不经稀释,即可用于检测。液体制剂混匀后静置2min即可消除气泡,冻干制剂则需静置2~4h方可脱气。10μm粒子的回收率为98.5%,25μm粒子的回收率为100.1%。共检测人血白蛋白130批、静注人免疫球蛋白（pH 4）103批、冻干静注人免疫球蛋白（pH 4）44批,均符合规定。结论《中国药典》2010年版三部附录中的＂不溶性微粒检查法＂可用于人血白蛋白、静注人免疫球蛋白的检测,抽检的样品均能达到新版药典的要求。     

三株不同细胞用于人干扰素效价测定的研究

本文采用细胞病变抑制法(微量法),用人羊膜细胞WISH株、FL株及人上皮细胞HEP2对3批精制人白细胞干扰素及12批粗制人白细胞干扰素进行了效价的检测,以中国药品生物制品检定所使用的测定细胞人羊膜细胞WISH株为标准,将HEP-2及FL细胞所测结果与其进行比较.经统计学处理,P>0.5,表明HEP-2及FL与WISH细胞测定结果没有差异,可用于人干扰素的效价测定.实验选用了EDTA作为细胞的消化剂,可高压灭菌、配制方便,适用于生产检测.     

人血白细胞制备α-干扰素转移因子和免疫核糖核酸的初步联产试验

报道一种利用同一批人外周血白细胞分别制备α-干扰素、转移因子和免疫核糖核酸的联产试验,可获得相同的α-干扰素产率及相应量的转移因子和免疫核糖核酸。     

干扰素来源的现状和展望

研究干扰素人员往往不能获得他们所需数量的干扰素用于研究,特别是用于临床研究。如果他们所进行的临床试验继续获得阳性结果,那么对干扰素的需求就会大大增加。迄今,临床试验用的所有人干扰素都是白细胞干扰素,它是从全血分离制备血浆、红细胞和其它医用物质时所得白细胞层制备的。这些白细胞很少有其它用处,但可储存,     

正交试验优化注射用重组人白细胞介素12处方

目的:优化注射用重组人白细胞介素12的处方。方法:采用正交试验设计,以甘露醇含量(A)、人血白蛋白含量(B)和pH值(C)为因素,活性、水分和外观的综合评分为指标优化处方,制备3批优化处方制剂并测定其活性、水分、pH值等。结果:优化处方为A4%、B0.5%、C7.4;3批制剂冻干后活性约为8400、8111、8511u.μg-1,水分<3%,pH值约为7.35～7.42,均符合质量标准要求(批间比较P>0.05)。结论:以优化处方制备的注射用重组人白细胞介素12活性较高,工艺重现性好。     

先锋必(注射用头孢哌酮钠)热原检查法和细菌内毒素检查法的比较性研究

本文阐述了应用热原检查法和细菌内毒素检査法同时对120批先锋必(注射用头孢哌酮钠)样品进行检查,并设立多项对比试验,以确定细菌内毒素检查法取代先锋必热原检查的可行性。结果发现,120批先锋必样品经热原检查和细菌内毒素检查均合格,样品内毒素含量远低于美国药典限值。通过测定加有一定量细菌内毒素的先锋必样品组及对照组的热原反应和鲎试验反应发现,在细菌内毒素浓度为10EU/Mし时,対照组可引发很强的热原反应,而先锋必样品(100MG/ML/KG)组所引发的热原反应明显减弱。说明先锋必在高浓度对热原反应有明显抑制作用。同样,本试验也发现,先锋必在较髙浓度对鲎试验也有明显抑制作用,其最高非抑制浓度为2.5MG/ML。对先锋必内毒素限值测定結果发现,按中国药典兔剂量注射,先锋必内毒素含量达0.1EU/MG时即可引发热原反应,而在0.2EU/MG时热原反应明显。本研究证实,细菌内毒素检査法可以代替热原检查法用于先锋必(注射用头孢哌酮钠)检查,中国药典90版规定的注射用头孢哌酮钠热原检查的兔剂M偏高,对热原反应有抑制作用。     

Gene delivery of albumin binding peptide-interferon-gamma fusion protein with improved pharmacokinetic properties and sustained biological activity

We have demonstrated that gene delivery of a fusion protein of mouse interferon (IFN) γ with mouse serum albumin (IFNγ–MSA) was effective in prolonging the circulation half-life of IFNγ in mice. However, the fusion to MSA greatly reduced the biological activity of IFNγ to less than 1%. In this study, we designed IFNγ fusion proteins with a 20 amino-acid long albumin-binding peptide (ABP) to prolong the in vivo half-life of IFNγ without reducing its biological activity. IFNγ–ABP and ABP–IFNγ, two fusion proteins with the ABP being fused to the C- or N-terminal of IFNγ, retained 40%–50% biological activities determined using a gamma-activated sequence-dependent luciferase assay. These fusion proteins exhibited the ability to bind to MSA. Gene delivery of IFNγ–ABP or ABP–IFNγ to mice using the hydrodynamic injection method resulted in a sustained concentration of IFNγ in the serum compared with gene delivery of IFNγ. In addition, the growth of mouse colon carcinoma CT-26 cells in the lung was efficiently inhibited by gene delivery of the IFNγ fusion proteins. These results indicate that the fusion of ABP is a useful approach to achieving prolonged retention in the blood circulation through binding to serum albumin and retaining biological activity.     

血清抗风疹IgG抗体检测方法的比较研究

目的  比较微量血凝抑制 （micro-hemagglutination inhibition,HI）试验与酶联免疫吸附测定（enzyme-linked immunosorbent assay,ELISA）检测血清抗风疹IgG抗体的敏感性和特异性,确定ELISA法检测血清抗风疹IgG抗体的可行性。方法  采集江苏地区506名8~15月龄健康儿童免疫前和麻疹-腮腺炎-风疹联合疫苗免疫后1个月的血清,共1012份血清,分别采用HI法和ELISA法检测血清抗风疹IgG抗体,并采用卡方检验对两种方法的检测结果进行比较。结果  HI法和ELISA法的血清抗风疹抗体阳性检出率分别为50.30%和47.33%,两种方法的阳性检出率之间的差异无统计学意义（χ²=1.780,P=0.182）。两种方法检出同为阴性的血清495份,同为阳性的血清476份血清,两种方法的检测符合率为95.95%。相关与回归分析显示,两种方法具有高度的正相关性（r=0.806,P＜0.001）。结论  可用ELISA法替代HI法进行血清抗风疹IgG抗体检测。     

127例乙肝表面抗体ELISA检测与化学发光法的比较

目的 探讨浓度处于5～100mIU/ml的乙肝表面抗体(HBsAb)ELISA定性检测与化学发光法的符合情况.方法 采用10种国产主流HBsAb ELISA试剂对127例经用化学发光法检测已知HBsAb浓度的标本进行检测,分析ELISA检测OD值与抗体浓度的相关性,并以试剂盒临界值为准分别比较不同OD值段ELISA结果 与化学发光法结果 的符合率.结果 10种同产HBsAb ELISA试剂盒OD值与化学发光定最值均呈正相关(P<0.01);ELISA法检测结果 与化学发光的阴阳符合率比较显示,5～10mIU/ml样本符合率较低,只有37.5%,在大于100mIU/ml时可达99.4%,在10～30mIU/ml,30～100mIU/ml分别为75.3%和90.7%;OD值存0～0.05,0.05～0.105、0.105～0.21、0.21～0.3、0.3～0.5、大于0.5分别为73.4%、65.3%、60.5%、72.0%、86.5%、92.2%.结果 显示阴阳结果 符合率在较高在大于0.5时最高,在介于0.105～0.21的符合率最低;处于试剂盒临界值±20%处的标本的符合率普遍偏低,最低只有42%,最高亦不超过80%.结论 虽然10种国产ELISA试剂之间的检测结果 没有统计学差异,但是对于浓度介于5～10mIU/ml标本符合率低.而且在接近试剂盒临界值时与化学发光结果 符合率则很低,但是随着OD值的增高符合率也逐渐升高.考虑到乙肝表面抗体的特殊临床意义,如何选取一个更为科学客观的结果 判断临界值值得进一步探讨.     

一种用鲎试验测定灭活内毒素的新方法(英文)

本文建立了一种用鲎试验测定灭活内毒素的新方法,其主要步骤包括:(1)样品与内毒素的预保温反应;(2)反应混合物的有限稀释;(3)用ELISA鲎试验测定残留的内毒素,样品灭活内毒素的活性(EIA50)定义为能灭活加入的内毒素的50％时的初始内毒素浓度。采用这种方法,可以观察到人血清和血浆,鲎血浆,鲎阿米巴细胞溶解物(LAL)以及多粘菌素B对内毒素的定量灭活作用。LAL对不同菌株的革兰氏阴性菌产生的内毒素的灭活作用有显著不同。本文建立的方法可以消除样品对鲎试验的干扰,适用于筛选和评价抗内毒素物质。     

The catalytic effect of bovine serum albumin on the ortho rearrangement of the potential ultimate carcinogen, N-(sulfooxy)-2-(acetylamino)fluorene, generated enzymatically from N-hydroxy-2-(acetylamino)fluorene and evidence for substrate specificity of the enzymatic sulfonation of arylhydroxamic acids.

This investigation examines the catalytic effect of bovine serum albumin on the ortho rearrangement of the possible ultimate carcinogen, N-(sulfooxy)-2-(acetylamino)fluorene, generated from N-hydroxy-2-(acetylamino)fluorene by the sulfotransferase(s) in the cytosol of rat liver. With various preparations of cytosol, 55-75% of the substrate, N-hydroxy-2-(acetylamino)-fluorene, was found to rearrange to the nonmutagenic and noncarcinogenic o-(sulfooxy) esters, 1- and 3-(sulfooxy)-2-(acetylamino)fluorene, in the presence of bovine serum albumin, while less than 1% of the substrate rearranged in its absence. In presence of bovine serum albumin the cytosolic reduction of N-(sulfooxy)-2-(acetylamino)fluorene to 2-(acetylamino)fluorene decreased by 60-90% and its solvolytic degradation to 4-hydroxy-2-(acetylamino)fluorene by 80-90%. The covalent interaction of enzymatically generated N-(sulfooxy)-2-(acetylamino)fluorene with the nucleophilic acceptors, N-acetyl-L-methionine and guanosine, was lowered by greater than 90% by addition of bovine serum albumin. These measurements indicated that the albumin-catalyzed ortho rearrangement controls the rates of concurrent metabolic and degradative reactions of N-(sulfooxy)-2-(acetylamino)fluorene. The results are in agreement with previous findings of a catalytic effect of serum albumin on the ortho rearrangement of synthetic N-(sulfooxy)-2-(acetylamino)fluorene. In contrast to its catalytic effect on the formation of o-(sulfooxy) esters from N-(sulfooxy)-2-(acetylamino)fluorene, bovine serum albumin had no effect on the formation of o-(acetylamino)fluorenols. To assess the substrate specificity of bovine serum albumin, its effect on the rearrangement of N-hydroxy-2-(benzoylamino)fluorene, a carcinogenic analogue of N-hydroxy-2-(acetylamino)fluorene, was analyzed under conditions of cytosolic sulfonation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Diazepam-sodium salicylate solution: Dilution with intravenous fluids,in vitro haemolytic activity and protein binding

The compatibility and stability of diazepam 5 mg/ml in 30% sodium salicylate, following dilution with 5% dextrose and normal saline, was investigated. Test dilutions ranging from 1 : 1 to 1 : 100 did not result in immediate precipitation and remained clear for at least 1h and 3h after dilution with 5% dextrose, normal saline or human plasma respectively. However, microcrystal precipitation was noted in some solutions thereafter.No precipitation was observed when diazepam-sodium salicylate solution was injected at varying rates into the tubing of 5% dextrose and saline infusions moving at low rates.Diazepam-sodium salicylate solution induced a higher degree of haemolysis in vitro and was less bound to bovine serum albumin than a commercial diazepam injection. Sodium salicylate competitively inhibited the binding of diazepam to bovine serum albumin.Further studies are necessary for the clinical evaluation of this diazepam-sodium salicylate combination.     

The lymphatic route--III. Pharmacokinetics of human natural interferon-beta injected with albumin as a retarder in rabbits

The aim of the present investigation was to define whether multisite, subcutaneous (s.c.) administration in unanesthetized, unrestrained rabbits of human natural interferon-beta (nat. IFN-beta) either in saline, or in a human albumin (ALB) solution (10 and 13% final concentrations) modified the pharmacokinetic parameters calculated from the IFN plasma levels. Plasma disappearance rates of nat. INF-beta were measured in two rabbits after intravenous (i.v.) administration and the kinetic was adequately represented by a bi-exponential curve. The highest ALB concentration (13%) caused a significant reduction of the plasma IFN Cmax, a longer half-life, a three-fold increase of the area under curve (AUC value) and a marked decrease of the plasma clearance. Interestingly, the bio-availability of IFN was increased almost four-fold. The data suggest that, when nat. IFN-beta is injected subcutaneously, the presence of a high concentration of ALB may prevent its inactivation and may favour its absorption via lymphatics rather than blood capillaries. It is remarkable that by using this approach, low but constant IFN levels are maintained for as long as two days, a fact that may well increase the therapeutic index of IFN in patients.     

Evaluation of quantification methods of occupational endotoxin exposure

Endotoxin has been identified as important component of organic-dust exposure and is suspected as main cause of work-related adverse health effects in dusty areas. Although the determination of endotoxin levels by using the Limulus amoebocyte lysate (LAL) assay is internationally accepted, reliability and variation of values measured with this test remain a point of discussion. Therefore, the purpose of the study was to determine the influence of different parameters on endotoxin activity measured in airborne samples. This study thus analyzed: (a) dust filter extraction procedures, (b) storage of samples, (c) usage of different commercially available LAL assays, and (d) results of the whole blood assay (WBA) compared to the LAL test. Using a parallel sampler, 120 filters were loaded with dust at 4 different occupational settings and extracted in 2 labs using a standardized protocol. Parameters like Tween in the extraction medium, extraction volume, centrifugation speed, and material of tubes used for extraction were tested. The LAL test and the WBA were able to determine the differences in dust load of filters obtained from the settings investigated. In addition, results varied significantly with modifications in extraction procedures. Using Tween for filter extraction mainly influenced the resulting endotoxin activity. In addition, LAL test differences according to manufacturer of LAL test, extraction volume, and whether the samples are freshly processed or frozen also resulted in significant variations in the endotoxin levels. In conclusion, a reliable assessment of exposure to endotoxin activity is only possible if standard operation procedures (SOPs) for sampling and determination are established.     

Prolonged circulation half-life of interferon γ activity by gene delivery of interferon γ-serum albumin fusion protein in mice

Gene delivery of mouse interferon (IFN) γ has been shown to inhibit metastatic tumor growth and onset of atopic dermatitis in mouse models. In this study, we tried to increase the circulation half-life of IFNγ after its gene delivery by designing a novel fusion protein of IFNγ with mouse serum albumin (MSA). Western blot analysis confirmed that IFNγ-MSA was expressed as a fusion protein, but hardly formed dimer as IFNγ did. The biological activity of IFNγ-MSA, which was examined using a plasmid expressing luciferase under the control of gamma-activated sequence elements, was about 200-fold lower than the activity of IFNγ. Intravenous injection of the proteins into mice confirmed that the circulation half-life of IFNγ was significantly prolonged by the modification. A hydrodynamic injection of a plasmid expressing IFNγ-MSA resulted in a sustained concentration in mouse serum; it resulted in about sixfold greater area under the concentration-time curve and about threefold longer mean residence time of IFNγ activity than those of IFNγ. Gene delivery of IFNγ-MSA inhibited tumor metastasis to a similar level to that of IFNγ despite the reduced activity of IFNγ-MSA. These results indicate that gene delivery of IFNγ-MSA is a promising approach to prolong the circulation half-life of IFNγ activity.