抗A、抗B血型单克隆抗体试剂稳定剂的筛选研究

临床上使用的抗A、抗B血型定型试剂根据其原材料来源,可分为多克隆抗体(免疫马血清)和单克隆抗体两大类,与多克隆抗体血型试剂相比,单抗血型试剂具有生产成本低、亲合力高等优点,但单抗血型试剂的稳定性欠佳。通过向单抗血型试剂中加入不同配比的稳定剂,观察其对抗A、抗B血型单抗     

国产人ABO血型定型试剂稳定性观察

为了解国内人ＡＢＯ血型定型试剂的稳定性,随机对４个厂家生产的共２０批次的血型试剂置２－８℃条件下,定期进行稳定性观察,结果显示,所有试剂在其效期内,凝集效价均下降１－２个滴度,其中有二个批次的试剂在临近失效期时,效价已低于１：１２８,其余全部合格,合格率为９０％,说明目前国产人ＡＢＯ血型定型试剂的稳定性是良好的,同时提示该剂的稳定性还需进一步提高。     

血型单克隆抗体试剂规模化生产工艺的建立

为了满足规模化生产的需要 ,对血型杂交瘤细胞的接种量、培养时间、培养条件等重要步骤进行了优化比较。结果表明 ,最佳血型细胞的接种量为 2 4× 10 8个 ,过长上清的凝集效价达到 1∶12 8,重复性好。建立了切实可行的血型试剂大规模生产工艺 ,成品试剂的质量稳定可靠     

尿微量白蛋白ELISA诊断试剂盒稳定剂效果研究

目的:观察尿微量白蛋白诊断ELISA试剂包被稳定剂和酶结合稳定剂的效果。方法:定期检测分别保存于37℃和4℃的尿微量白蛋白试剂盒,并对其稳定性进行评估。结果:该尿微量白蛋白诊断试剂包被稳定剂和酶结合稳定剂可以使尿微量白蛋白试剂盒在2~8℃条件下放置6个月以上。     

人红细胞非凝集型单抗(自身全血凝集试验试剂)的研制及鉴定

应用杂交瘤技术 ,混合人O型红细胞免疫纯系BALB/C小鼠 ,包被的红细胞作为固相抗原 ,ELISA法筛选出识别红细胞表面共同抗原的抗体 ,然后利用Coombs原理 ,加入抗球蛋白试剂。用玻片法筛选出非凝集型抗体 ,得到 2株稳定分泌上述抗体的杂交瘤并进行初步鉴定 ,单抗上清血凝效价可达 1∶5 0～ 1∶10 0 ,腹水血凝效价为 1∶8× 10 4 ～ 1∶16× 10 4 。另外 ,本文特别讨论了凝集型单抗的Fab段作为非凝集型抗体应用的可能性。     

抗血型物质杂交瘤细胞悬浮培养研究

诱导、选育了适于连续悬浮培养的抗人血型杂交瘤细胞株,并初步建立了一种血型定型试剂悬浮培养生产工艺,抗体效价和亲和力等所有指标均达到或超过国家标准。它是一种经济、简单、切实可行的大生产方法。此外,为用发酵罐大规模生产血型定型试剂提供了物质基础和初步的生产模式。     

马免疫血浆的稳定性研究

目的了解不同温度条件对马免疫血浆外观和效价的稳定性影响,为马免疫血浆的采集、分离、贮存、运输及抗毒素生产提供数据支持。方法将破伤风类毒素及肉毒A、B、E、F型类毒素制备的马免疫血浆,分别放置于2~8℃、20℃、37℃3种温度下,并分别于0、1、3、6、12个月取样,依据《中国药典》三部(2010版)的检测方法和标准,对样品进行外观检查及效价检测。结果破伤风及肉毒A、B、E、F型马免疫血浆在2~8℃条件下,放置12个月稳定性良好,外观及效价均符合《中国药典》三部(2010版)规定要求。20℃与37℃下放置的马免疫血浆随着时间的延长,外观会发生不同程度的浑浊,效价也均有不同程度的降低,低效价组比中、高效价组的效价下降明显,且温度越高效价降低幅度越大。结论马免疫血浆在2~8℃条件下质量稳定。     

人ABO血型定型试剂(单克隆抗体)临床试验

通过与１１家单位协作,对中国药品生物制品检定所分发的ＡＢＯ血型单克隆抗体定型试剂进行临床考核,检测１～１２个月婴儿、５９岁以上老人、１～５９岁成人、健康献血员、癌症患者及其它疾病患者共计７０９６８例,单抗试剂检测结果与反定型结果完全相符,特别是对红细胞表面抗原位点少、亚型抗原性弱的新生儿和红细胞表面抗原成份可能发生变化的癌症患者,均呈现较好的凝集模式。     

血型检测用反向定型试剂的研制

将正常的红细胞在特定条件下用甲醛处理,使红细胞膜固定但不影响膜表面糖蛋白血型抗原的活性。采用与正向定型相同的平板凝集试验方法,４０６０份血样正向和反向定型结果完全一致。经稳定性观察９０天,处理后的红细胞与相应抗体的凝集性能未见明显改变。实验结果表明本文介绍的红细胞试剂可用于ＡＢＯ血型鉴定的反向定型试验。     

冻干人用狂犬病疫苗(Vero细胞)稳定剂配方的筛选

目的筛选出适合冻干人用狂犬病疫苗(Vero细胞)的热稳定性好、无明胶的稳定剂配方。方法以水分、外观、效价、热稳定性效价为指标,尤其是效价和热稳定性效价,对A、B、C、D、E、F、G等6种稳定剂配方进行系统的优化组合筛选,6种稳定剂配方分别含有蔗糖、乳糖、人血白蛋白、甘氨酸、精氨酸、明胶、尿素、甘露醇、右旋糖苷,其中A为现行疫苗生产配方。运用单因素五元设计法,筛选出最优稳定剂组合。结果方差分析结果显示D组配方对冻干人用狂犬病疫苗(Vero细胞)的M-ABT结果 13.95,37℃放置4周M-ABT结果 13.05;NIH效价1.94,37℃放置4周NIH效价1.56。结论 D组配方对冻干人用狂犬病疫苗(Vero细胞)有较好的保护作用。     

Rare B(A)02/O01 was found in Sichuan again: a case report

B(A) is a rare ABO blood subgroup. Here we reported a B(A)02/O01 case. One 25-year-old female patient showed inconsistent forward and reverse blood grouping results based on micro-column gel agglutination assay. PCR-SSP and PCR-SBT based genotyping indicated that the patient was B(A)02/O01 heterozygous.     

Dried Escherichia diagnostic agents from live cultures

The possibility of using selected cultures of opportunistic Escherichia, freeze-dried in a saccharose-gelatine medium, as dried diagnostic reagents. In the agglutination test with Escherichia OK diagnostic sera, both fresh and stored for a period of up to 1 year, these dried diagnostic reagents prepared from live cultures showed the same degree of agglutination as live suspensions of similar Escherichia cultures.     

Effect of vitamin A analogs on concanavalin A-induced agglutination of erythrocytes

The vitamin A analogs retinoic acid and retinol caused a significant increase in concanavalin A-induced agglutination of human erythrocytes, while its esters, retinyl acetate and retinyl palmitate, were found to be ineffective. The effect of membrane labilizers and stabilizers on the enhancement of agglutination as well as the properties of the model system employed showed that the action of vitamin A is due to a direct action on cell membrane and is not mediated by the release of lysosomal proteases into the medium, a hypothesis proposed by earlier workers.     

Mixture of three amino acids as stabilizers replacing albumin in lyophilization of new third generation recombinant factor VIII GreenGene F

A formulation with stabilizers replacing albumin was developed for lyophilization of recombinant factor VIII (FVIII), GreenGene F (WHO INN: beroctocog alfa), to achieve stability and eliminate safety issues of blood‐derived albumin. L ‐Arginine (hydrophilic amino acid, positively charged side chain), L ‐glutamic acid (hydrophilic amino acid, negatively charged side chain), and L ‐isoleucine (hydrophobic amino acid, nonpolar) were selected as stabilizers, and the mixture of the three amino acids were optimized. The mixture had results comparative with albumin and other commonly used stabilizers showing good preservation of recombinant FVIII during lyophilization, robust stability with consistently high recovery of FVIIII, very low aggregate formation, and good storage stability without alterations in protein characteristics. In vivo test results showed that the efficacy was maintained and had no signs of toxicity. The study demonstrated that the three amino acid mixture acts as a good stabilizer for lyophilization of recombinant FVIII and as a safe excipient. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

Turbidimetric study of latex agglutination in 2,4-dichlorophenoxyacetic acid analysis

By the example of 2,4-dichlorophenoxyacetic acid (2,4-D) assay by the inhibition of latex agglutination, new synthetic polymer microspheres for the conjugation with antibodies to 2,4-D and agglutinators based on ovalbumin and polyacrylamide were developed and characterized. The effect of various parameters (the concentration of reagents, the type and the degree of modification of the microsphere surface, and the nature of the carrier in the composition of the agglutinator) on the rate of agglutination and the maximal optical absorption observed during the reaction were studied by turbidimetry. The optimal parameters were found for the assay of 2,4-D by the inhibition of latex agglutination with turbidimetric registration of the results.     

The turbidimetric study of latex particle agglutination: Assay of 2,4-dichlorophenoxyacetic acid

By the example of 2,4-dichlorophenoxyacetic acid (2,4-D) assay by the inhibition of latex agglutination, new synthetic polymer microspheres for the conjugation with antibodies to 2,4-D and agglutinators based on ovalbumin and polyacrylamide were developed and characterized. The effect of various parameters (the concentration of reagents, the type and the degree of modification of the microsphere surface, and the nature of the carrier in the composition of the agglutinator) on the rate of agglutination and the maximal optical absorption observed during the reaction were studied by turbidimetry. The optimal parameters were found for the assay of 2,4-D by the inhibition of latex agglutination with turbidimetric registration of the results.     

Serological and molecular analysis of ABO and Rh blood group chimeras

The serological examination, blood transfusion strategies and the molecular analysis to blood group chimera were conducted to demonstrate existent of chimera in blood group. The blood grouping of ABO or/and RhD, newborn red blood cells separated by capillary centrifugation. Aabsorption tests and DTT treated agglutination erythrocyte tests were implemented in four patients. Further molecular biological research was conducted on one patient''s sample. The results showed that for patient 1: ABO blood group was AB/B chimera, Rh blood cells contained the RhCE chimera gene; Patient 2: Rh blood cells contained the RhD chimera gene; Patient 3: ABO blood group was AB/B chimera, Rh blood cells contained the RhD chimera gene; Patient 4: ABO blood group was O/B chimera, Rh blood cells contained the RhCE chimera gene. The study suggests that the individuals categorized as chimeras are likely to be more common than existing literature reports. According to the serological tests, in the absence of a history of recent blood transfusion or disease to cause reduced antigen, the phenomena of hybrid aggregation of the ABO and Rh blood system were the main feature. In terms of transfusion strategy, the selection of ABO and Rh blood groups should be depended on the group of cells with more antigens.     

绿僵菌属血清学的初步研究

本文采用凝集试验和免疫电泳的方法对不同来源的七株绿僵菌属真菌进行了免疫学对比研究。抗原是孢子悬液和菌丝体清液,通过对家兔接种抗原而获得抗血清。每一种抗原与其同源抗血清和异源抗血清进行交叉试验,并对凝集试验的结果以及免疫电泳反应产生的沉淀弧数目和免疫电泳图谱加以对比分析,试验结果表明:不同菌株间存在着明显的抗原类似性,同时各菌株间也表现出一定的抗原专一性。根据试验数据对供试菌株进行血清学分型的结果与形态学分类相符合。     

The standardization of the candidate national standard anti-D blood-typing serum with the Groupamatic MG50 system

The utility of the Groupamatic MG50 system for quantitative expression of agglutination in terms of continuous response was studied. By use of the characteristics of the design of this system, in which the change in voltage reflects the degree of agglutination, a linear regression relationship between the log ratio of light flux obtained by transformation of the voltage and the log dilution factor of the serum was demonstrated. The availability of the parallel line assay method to the standardization of the blood grouping antisera was also described.     

南方鲇血型的初步鉴定

本研究旨在通过观察南方鲇血清与其红细胞的交叉反应以鉴定南方鲇的血型.实验结果表明:南方鲇的血清与同种其他个体的红细胞进行交叉反应时均未出现凝集现象,这表明南方鲇可能不存在血型或南方鲇具备血型但血清中相应的凝集素含量不足.以南方鲇的红细胞为抗原免疫日本种大耳白兔制备的抗血清与南方鲇的红细胞进行交叉反应,出现了不同程度的凝集反应,这表明南方鲇存在血型.据上述两个实验结果可以推断,南方鲇可能存在4种血型,分别命名为NA、NB、NAB和NO型;同时也证实,在鉴定南方鲇血型的研究中,通过制备抗血清与红细胞进行交叉反应的方法更为可靠.