卡式微柱凝胶法在猴血型鉴定中的应用

灵长类动物的ABO血型抗原都表达在组织器官内,而不是在红细胞上,这给灵长类动物血型的鉴定带来很大的困难。为找到更加简捷、准确鉴定灵长类动物类人ABO血型的方法,采用近年来临床上广泛应用的卡式微柱凝胶正、反定型法对34只猕猴和16只食蟹猴的血型进行了鉴定,并与肾组织免疫组化法的检测结果进行比较。结果显示:卡式微柱凝胶正定型法的检测结果中无一例为阳性结果;血浆中的纤维蛋白原和人-猴种属间非特异性抗体都会对卡式微柱凝胶反定型法的部分检测结果产生干扰;采用经正常人O型红细胞吸附处理后的清亮血清,卡式微柱凝胶反定型法的检测结果明确,与免疫组化法判定结果一致。由此得出:卡式微柱凝胶反定型法可以用于灵长类动物血型的鉴定,其主要干扰因素为血浆内的纤维蛋白原和人-猴种属间非特异性抗体,在采用清亮血清及经正常人O型红细胞吸附处理后能消除其干扰。     

自身抗体引起的ABO血型正反定型结果不一致

目的:对因自身抗体引起ABO血型正反定型结果不一致的疑难血标本进行血型鉴定和分析,探讨其原因和解决办法。方法:先用直接抗人球蛋白试验(DAT)和间接抗人球蛋白试验(IAT)确定ABO血型正反定型结果不一致的原因是由于自身抗体的存在,然后做吸收放散试验、抗体筛选试验等排除自身抗体的干扰以便血型的正确判定。结果:ABO血型正反定型结果不一致因自身抗体引起的有10例,其中温抗体7例,冷抗体3例,温抗体同时同种抗体阳性2例,冷抗体同时同种抗体阳性2例。结论:自身抗体会影响ABO血型的正确判定,选用合适的试验进行分析判断以提高血型鉴定的准确性。     

ABO血型正反定型及交叉配血实验在外科病人手术输血中应用

目的:探讨ABO血型正反定型及交叉配血实验在外科手术患者输血中的应用效果及影响因素。方法:选取我院自2017年2月-2019年2月收治的80例行ABO正反定型与交叉配血治疗的外科手术患者,记录ABO反定型与交叉配血不合的标本,使用2-Me处理被患者自身冷抗体凝集的红细胞,同时使用微柱凝胶法、凝聚胺法对血型不规则抗体以及特异性进行筛选和鉴定。分析ABO血型反定型不符合以及交叉配血不合的影响因素。结果:对正反定型完全无凝集反应的80例血清标本进行交叉配血实验,其中8例存在凝集反应,配血不合情况;导致外科手术患者输血中ABO血型反定型不符交叉配血不合的主要因素包括自身冷抗体、血型抗原性减弱、血型不规则抗体以及血型抗体效价减弱等。结论:ABO血型正反定型及交叉配血治疗中的患者中,大部分配血一致,少数的交叉配血不合,主要与自身冷抗体、血型抗原性减弱、血型不规则抗体以及血型抗体效价减弱等因素相关。     

全自动血型仪的应用评价

目的了解全自动血型鉴定系统用于检测ABO血型和Rh(D)血型的效果。方法采用深圳爱康AK03A型数字血型仪对北海市中心血站2010年5月—2011年12月无偿献血者20702份样本进行ABO血型和Rh(D)血型鉴定,并与手工试管法进行对比验证。结果发现正反定型不一致的有75例,ABO初筛血型错误的36例,Rh(D)阴性的53例。结论全自动血型鉴定系统检测血型快捷、方便、高效,能降低检测人员的工作强度,但判读结果时需要与手工试管法结合进行验证,才能得出正确的结果。     

1939例ABO血型系统新生儿溶血病的血型分布

目的：探讨在母婴ABO血型不合引起的新生儿溶血病（HDN）中的血型分布。方法：对临床表现怀疑为HDN的1939例新生儿进行血清学试验检测,包括新生儿红细胞直接抗人球蛋白试验,红细胞抗体释放试验和游离抗体试验,同时检测母亲ABO血型和RhD血型。结果：1939例婴儿母亲血型均为O型RhD阳性。A型血新生儿818例,其中直接抗人球蛋白试验阳性率17.2%（141/818）,红细胞抗体释放试验阳性率89.2%（730/818）。B型血婴儿1121例,其中直接抗人球蛋白试验阳性率10.9%（121/1121）,红细胞抗体释放试验阳性率78.6%（881/1121）。结论：ABO血型不合引起的HDN中,A型新生儿比B型新生儿患病几率更大。     

Bioxun微柱凝胶筛查红细胞不规则抗体的研究

目的:探讨Bioxun微柱凝胶法(Bioxun Micro-column Gel anti-globulin Test,BMGT)筛查红细胞不规则抗体的敏感性和特异性。方法:使用盐水介质法(Saline Media Test,SMT)、传统抗人球法(Traditional anti-Globulin Test,TGT)、达亚美凝胶法(DiamedMicro-column Gel Test,DMGT)和Bioxun微柱凝胶法分别检测红细胞不规则抗体,对方法学灵敏度和特异性进行观察比较。结果:Bioxun微柱凝胶法与达亚美凝胶法的结果无显著差异(P>0.05),二者的敏感度和特异性超过盐水介质法和传统抗人球法(P<0.05)。结论:Bioxun微柱凝胶法操作快捷,敏感度高,适宜在临床输血中推广应用。     

卡式微柱凝胶法在ABO、RhD血型鉴定中的应用价值

目的:探讨微柱法在ABO、RhD血型鉴定中的应用价值。方法:抽取2012年3月至2013年3月我站检验科血液筛查标本共1081份作为研究对象,在metis全自动血型仪进行初筛的基础上,同时进行卡式微柱凝胶法ABO、RhD血型鉴定,据定型结果配血,统计输血结局并作为评价依据。结果:筛查RhD阴性者6份占0.56%,阳性者1075份占99.44%;所有血液输用后均未产生输血反应。结论:ABO、RhD血型鉴定是保障输血安全、抑制溶血性疾病的必要措施,而微柱法具有操作简便、价格低廉、科学准确的优点,适合血站的血型鉴定和临床的推广。     

2623名患者红细胞不规则抗体筛选结果分析*

目的:对红细胞抗体筛选阳性的标本进行红细胞不规则抗体特异性鉴定与分析,为临床安全输血提供帮助。方法:2011年8月至2018年12月天津市各医院送至本中心实验室检测的2623份血液标本,进行红细胞不规则抗体筛选和鉴定;采用低离子强度溶液-抗人球蛋白微柱凝胶卡方法和盐水直接离心方法进行抗体筛选和抗体鉴定。结果:共检出2744个红细胞抗体,其中2139个为同种抗体,509个为自身抗体,96份标本未能确定抗体特异性;在所检出的同种抗体中,Rh系统的抗体最常见,占比为54.7％(其中抗-E 26.2％,抗-c 8.7％,抗-C 7.7％,抗-e 6.9％,抗-D 4.9％),MNS系统、Lewis系统、Kidd系统、P系统、Duffy系统、Kell系统、Diego系统均有抗体检出。结论:在输血前检测中,为预防检出不规则抗体的情况出现,应提前备血,以免由此导致的延迟输血,影响治疗。     

应用SELEX技术筛选人类血型IgG适体的实验研究

目的:建立Rh(D)血型的IgG类抗体适体的筛选方法,为后续合成适体并进行新生儿溶血病的防治研究奠定基础。方法:利用SELEX技术,构建含有52个随机序列的单链DNA文库,利用硝酸纤维素膜法筛选与IgG类抗体分子高亲和力结合的核酸分子,同时探索并优化将其扩增为双链DNA核酸库的PCR反应条件;通过膜结合实验检测核酸分子的富集效果并用凝胶阻滞实验初步测定所得核酸适体与Rh(D)血型IgG类抗体的亲和力。结果:随着筛选的进行,核酸分子的富集库向着与靶分子亲和性增强的方向进化,经过了11个循环,筛选出与Rh(D)血型IgG抗体结合力较强的核酸分子,与核酸结合的IgG分子在凝胶阻滞实验中显示出阻滞带。结论:初步建立了利用SELEX技术筛选人类Rh(D)血型IgG抗体适体的方法。     

抗-D抗体及其应用

Rh血型是仅次于ABO血型系统的人类红细胞抗原系统,至今已发现40多种抗原,但与临床密切相关的是D,C、c、E、e等5种抗原,其中最主要的是D抗原。相应的抗-D抗体无论是在临床输血检测,还是在Rh(D)新生儿溶血病、溶血性输血反应等的防治方面均具有非常重要的意义。传统的抗-D抗体的制备需用人的血清,来源受限。各种抗-D人源性单克隆抗体和基因工程抗体已经成为发展方向。     

Application of new enhanced medium in indirect antiglobulin test by gel method

The objective of this study was to explore the efficacy of a new enhanced medium in indirect antiglobulin test by optimized gel method. Anti-IgG+ C3d and anti-IgG antiglobulin gel cards, and samples from three blood group systems: Rh, Kidd and MNSs were used. The new enhanced medium was incubated at 37°C for 5 minutes, low ionic strength solution (LISS) for 15 minutes, saline solution for 30 minutes, respectively. The sensitivity and specificity of blood group antibodies were then compared. The results showed that the specificity and sensitivity of the new enhanced medium incubated for 5 minutes were consistent with those of LISS and saline solution for 15 minutes and 30 minutes, respectively. The results suggest that the new enhanced medium be able to accelerate the binding of erythrocyte antibodies with antigens, reduce incubation time and optimize the detection process of indirect antiglobulin test by gel method.     

Assessment and value of the polybrene microplate test for study and identification of irregular anti-erythrocyte antibodies

We have adapted Lalezari's manual polybrene test for use with microplate technology for screening and identification of anti-erythrocytes antibodies with a view to future automation. The technical conditions have been standardized, firstly by using a programmable centrifuge and a sequential shaking, secondly by using a preservative medium for panel after dispensing onto microplates. This methodology has been run in parallel with papain test and LIS indirect antiglobulin test: 7,000 screenings have been performed and their results are considered here. Our results are comparable to those described for automatic and manual techniques. The polybrene-microplate test affords a fast, reliable, handy and inexpensive means of screening and identification for irregular antibodies. It appears as an additional method for enzymatic tests in microplate. An antiglobulin test can be carried out after negative tests.     

Biochemistry of the erythrocyte Rh polypeptides: a review

The clinically important Rh blood group system is complex, consisting of multiple distinct antigens. Despite clinical recognition for over 50 years, the Rh blood group antigens have remained poorly understood on a molecular level until the recent identification and characterization of the "Rh polypeptides," the core structural proteins of the Rh antigens. This group of erythrocyte membrane proteins of molecular weight 30,000-35,000 daltons was first recognized by employing Rh-specific antibodies to immunoprecipitate radiolabeled components of erythrocyte membranes. By using antibodies specific for the Rh D, c, and E antigens, a series of highly related non-identical proteins were immunoprecipitated, indicating that the Rh antigens are composed of multiple related proteins. The Rh polypeptides have been purified and characterized, and they were found to have several unusual biochemical characteristics. The Rh polypeptides penetrate the membrane bilayer; they are linked to the underlying membrane skeleton; they are covalently fatty acid acylated with palmitate. While the Rh antigenic reactivity is unique to human erythrocytes, the Rh polypeptides have been isolated from erythrocytes of diverse species and are thought to be fundamental components of all mammalian erythrocyte membranes. The functional role of the Rh polypeptides remains undefined, but a role in the organization of membrane phospholipid is suspected.     

Heterogeneity of antibody response to Salmonella lipopolysaccharide measured by passive hemagglutination and hemolysis in mice

The complement-requiring passive hemolysis test with Salmonella typhimurium lipopolysaccharide-coated sheep erythrocytes is more sensitive for antibodies directed against the lipopolysaccharide than is the passive hemagglutination test. The hemagglutinating and hemolyzing antibodies produced in Swiss mice by hyperimmunization, either with or without Freund's adjuvant, were distributed in both the light and heavy fractions isolated by sucrose density gradient fractionation and gel filtration. IgM fractions, whether tested by hemagglutination or hemolysis, were sensitive to 2-mercaptoethanol (0.15 m). On the other hand, IgG hemolytic antibodies were more sensitive to 2-mercaptoethanol than were IgG hemagglutinating antibodies. The resistance of IgG hemagglutinating activity amounted to about 72 to 95% of the total IgG recovered, whereas the resistant portion of the IgG hemolytic activity was approximately 40 to 53%. It is suggested that, although mercaptoethanol sensitivity is not a definitive test for IgM antibody, its use in connection with the hemagglutination test gives at least an approximation of the IgG antibody, whereas the hemolysis test gives a better approximation of maximal measurable antibody against Salmonella lipopolysaccharides.     

The Prevalence of unexpected antibodies in Saudi's plasma prior blood transfusion and their association with clinical conditions: A cross-sectional study

Unexpected antibodies, also called irregular antibodies, are not known to exist in a person's serum before testing. This research aims to assess the prevalence of unexpected antibodies and their correlation with several clinical conditions. This cross-sectional prospective study, undertaken from June 2019 to June 2020, included ABO, Rh grouping, cross-matching, and antibody screening. Antibody identification was performed only on patients who tested positive in the screening test. From a total of 9764 participants who were screened for unexpected antibodies, 107 (1.1%) tested positive. The Rh blood group system antibodies were the most frequent, particularly anti-D. There was also a significant correlation between the unexpected antibodies and history of transfusion, pregnancy, and autoimmune diseases as P ≤ 0.05. The most prominent unexpected antibodies in the study belong to the Rh system (Anti-D). Moreover, as a result of the strong correlation between the unexpected antibodies as well as the history of transfusion, pregnancy, and autoimmune diseases, the highest safety criteria must be followed during the transfusion of blood to patients with these clinical conditions.     

BLOOD TRANSFUSIONS AND THE Rh FACTOR

When Rh-negative persons are given transfusions of Rh-positive blood, more than 50 per cent are sensitized to the Rh₀ factor. Such sensitization of female children may be the cause of hemolytic disease in their offspring many years later, while severe hemolytic reactions may follow a second transfusion of Rh-positive blood in either sex.The gross hemolysis of transfused blood may be entirely asymptomatic, however. In one case a pint of blood was completely hemolyzed within two hours without producing symptoms. The only signs were hemoglobinuria, low grade jaundice, urobilinogenuria and a rising Rh antibody titer. The patient had previously been sensitized by a single pint of Rh-positive blood.The dangers of Rh sensitization can be avoided by routine Rh typing of all prospective recipients of transfusion, whether male or female, and by giving only Rh-negative blood to those who are Rh-negative.     

Characterization of the Mouse Rh Blood Group Gene

The Rh blood group system is of clinical importance in blood transfusion and as the cause of hemolytic disease of the newborn. Other than their role as carriers of Rh antigens, very little is known about the function of the Rh polypeptides. As a first step to generate an animal model system in which to study the structure and function of Rh, and to extend the phylogenetic analysis of RH genes, the Rh homologue from Mus musculus was characterized. Comparison of RH from humans and mice revealed 71 and 58% sequence identity at the nucleotide and amino acid levels, respectively. Mouse Rh mRNA encodes a protein which is 1 amino acid longer (418 aa) than that of human (417 aa). Rh protein was detected in mouse erythrocyte membranes and was comparable in size to human Rh. Mouse erythrocytes do not show serologic reactivity with human Rh antibodies, probably because the greatest divergence between the mouse and the human genes was seen in the predicted extracellular loops, while the transmembrane regions were more conserved. The mouse RH locus consists of only one gene, which is important for future genetic manipulation and which also indicates that the RH gene duplication seen in humans has occurred since the mammalian radiation.     

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.     

HEMATOLOGIC FEATURES WHICH MAY INFLUENCE Rh IMMUNIZATION

The biologic features which may influence Rh sensitization in the mother and hemolytic disease in the newborn are conditioned by the hereditary immunologic pattern of the subjects, the antigenic characteristic of the blood group specific substances, the character of the antibody responses, the inherent susceptibility to disease as a result of the immune mechanism, and the character and sequence of exposure to the antigenic substance. In respect to Rh-negative women, there appears to be a greater vulnerability to immunization among those of blood group A. In regard to the infant, the compatibility of the maternal-infant blood types and the part they play in the phenomenon of the competition of antigens may influence the occurrence of hemolytic disease in the newborn.