Epidemiology and immunogenetic background of islet cell antibody--positive nondiabetic schoolchildren. Ulm-Frankfurt population study.

Islet cell antibodies (ICAs) were determined in a large cohort of white nondiabetic schoolchildren (n = 4287) from a homogenous population in southern Germany. The prevalence of ICA levels greater than or equal to 5 Juvenile Diabetes Foundation (JDF) U was 1.05% (95% confidence interval 0.8-1.4%). Analysis of HLA-DR beta and -DQ beta alleles revealed that the specificities found to be increased in insulin-dependent (type I) diabetic subjects with the same ethnic background were also associated with ICA positivity in the nondiabetic schoolchildren. HLA-DR3 (P less than 0.01) and -DR4 (P less than 0.01) phenotypes and absence of Asp residue (P less than 0.01) at codon 57 of the HLA-DQ beta-chain were significantly increased in ICA+ compared with control subjects. High levels of ICAs, which were categorized as either greater than or equal to 17 or greater than or equal to 30 JDF U, were found to be associated with amino acids other than Asp at position 57 of the HLA-DQ beta-chain. No association of ICA level was found for HLA-DR phenotypes.

A new second-generation anti-HIV-1 enzyme immunoassay using recombinant envelope and core proteins.

In a multicenter collaborative study a new second-generation HIV-1 antibody enzyme immunoassay (Abbott recombinant HIV-1 EIA) using Escherichia coli-expressed recombinant p24 and p41 proteins as solid-phase antigens was compared with the first-generation H9 cell-line-based Abbott HIV-1 EIA. The results of the confirmatory assays (Western blot, immunofluorescence), combined with clinical information, were used as the reference standard for the detection of HIV-1 antibodies in 10,676 random blood donor serum specimens, in a panel of 840 specimens from symptomatic and asymptomatic patients and a total of 63 serial blood specimens from 23 people at risk. With fresh blood donor sera, the specificity of the first-generation assay ranged between 99.54 and 99.76% (95% confidence limits, CL) compared with 99.81-99.95% (95% CL) for the second-generation EIA. With panel specimens the recombinant HIV-1 EIA achieved an overall sensitivity of 100% and a specificity range of 98.3-99.7% (95% CL); the corresponding sensitivity and specificity ranges observed for the first-generation EIA were 98.0-99.5% (95% CL) and 94.3-96.8% (95% CL), respectively. The improved sensitivity for the second-generation assay was confirmed by testing serial samples from seroconverting patients. The use of recombinant proteins eliminated non-specific reactions due to class II human leukocyte antigen (HLA)-directed antibodies.

A rapid and sensitive immunoassay for antibodies against alloantigens on human platelet glycoproteins (BIPA).

A rapid and sensitive enzyme immunoassay has been developed for the detection and specification of alloantibodies against human platelet antigens (HPA) on platelet membrane glycoproteins. Human platelet glycoprotein complexes were coupled by monoclonal antibodies to immunomagnetic beads with sheep anti-mouse IgG directed antibodies. These beads served as solid-phase target antigens for human alloantibodies in a modified MAIPA assay. This test provided high sensitivity, permitting the identification of antibodies against membrane receptors and required only 1 h of assay time. The specificity and sensitivity of this assay were evaluated in serum samples from polytransfused patients and compared with standard immunoassays employing antigen coated immunowells.

The HTLV type I sequence from an asymptomatic German blood donor is related to sequences from South America.

In most parts of Europe only a limited number of sporadic cases of HTLV-I infections have been identified. So far, the few cases found in Germany were individuals from endemic areas or with relations to endemic areas. Here we report an HTLV-I infection from an asymptomatic female German blood donor whose only known potential risk was a former partner from South America, where HTLV-I is known to be endemic. The DNA sequence of the LTR region was determined and a phylogenetic analysis indeed suggested homologies with HTLV-I sequences from South America.

Increase of mobilized CD34-positive peripheral blood progenitor cells in patients with Hodgkin's disease, non-Hodgkin's lymphoma, and cancer of the testis.

G-CSF (filgrastim) can effectively mobilize peripheral blood progenitor cells (PBPC) when administered during steady-state hematopoiesis. In this single center study, we compared the effectiveness of two different doses of G-CSF on the mobilization of peripheral blood stem cells in patients with Hodgkin's disease, non-Hodgkin's lymphoma, and cancer of the testis. A first group including 33 patients received 10 micrograms G-CSF/kg BW per day (group A), whereas a second group comprising 34 patients was treated with 24 (2 x 12) micrograms G-CSF/kg body weight (BW) per day (group B) prior to the leukapheresis. A significant difference (P = 0.015) in the total number of CD34+ cells between group A: 11.32 x 10(7) (range 0.34-110.2) and group B: 48.25 x 10(7) (range 1.33-447.4) has been observed in the first leukapheresis product. Moreover, the total number of CFU-GM increased significantly from 34.79 x 10(4) (range 1.07-300.9) to 147.69 x 10(4) (range 1.03- 1204.0) (P < 0.005), and the number of MNC increased from 1.35 x 10(10) (range 0.41-3.09) group A) to 2.93 x 10(10) (range 0.66-9.7) (group B) (P < 0.001). Comparable results were obtained in the second leukapheresis. Our data indicate, that the application of higher doses of G-CSF can significantly improve the effectiveness of mobilizing PBPC during steady-state conditions, and thereby considerably contribute to a safe and fast engraftment as well as a reduced number of leukapheresis procedures to achieve sufficient number of PBPC.

German consensus on immunogenetic donor search for transplantation of allogeneic bone marrow and peripheral blood stem cells.

In Germany allotransplantation of bone marrow or peripheral blood stem cells is presently performed by 34 different teams operating more or less independently. Thus, strategies of immunogenetic donor search, use of the various tissue typing techniques and policy on acceptable HLA mismatches in related and unrelated settings may vary considerably from one transplant centre to another. This paper summarises the results of the first German consensus meeting on immunogenetic donor search for bone marrow/peripheral blood stem cell grafting. The main goal of the participating transplant physicians and immunogeneticists was to define national standards for the above issues.

Influence of stent length and heparin coating on platelet activation: a flow cytometric analysis in a pulsed floating model.

Platelets are involved in acute and subacute thrombotic occlusions of coronary stents and also may play a role in the pathophysiology of in-stent restenosis. This study sought to investigate the expression of activation dependent glycoproteins on platelets by flow cytometry and time until stent thrombosis in an in vitro model of stent thrombosis. Coronary stents were placed in parallel silicon tubings with circulating citrated platelet rich plasma to measure 1) influence of stent length on platelet antigens; 2) influence of heparin coating on platelet antigens; and 3) time until stent thrombosis. After recalcification aliquots of platelet-rich plasma were taken over 10 minutes in 2-minute intervals and immediately fixed and stabilized. For flow cytometric analysis monoclonal antibodies to CD41a (glycoprotein IIb/ IIIa), CD42b (glycoprotein Ib-V-IX), CD62p (P-selectin), and CD63 (glycoprotein 53) were used. Within 2 minutes after start of circulation, the expression of CD62p and CD63 increased. Longer stents resulted in more platelet activation than shorter stents (25 mm vs. 15 mm; p<0.001. Time until stent thrombosis was reduced (25 mm vs. 15 mm; p<0.05). Heparin coating did not significantly influence flow cytometry detectable platelet activation but prolonged time until stent thrombosis (coated vs. uncoated; p<0.005). In control tubing systems without stents platelet activation was less pronounced (p<0.0001). Antibodies to CD41a and CD42b did not show significant changes. In this model platelet activation detected by flow cytometry and time until stent thrombosis were dependent on stent length and coating. In vitro testing could be useful to optimize stent design and material.

HLA and obesity.

A possible association between HLA and obesity was investigated in 42 unrelated obese children from Hessen, Germany. A family study was possible in 11 families with a total of 26 children. No significant deviation was observed for one of the HLA ABC antigen frequencies in the sample group compared with controls (n = 220). The linkage analysis in families: a) ruled out a close linkage (less than 10 cM) between HLA and a hypothetical obesity gene (assumption of a dominant mode of inheritance), b) did not exclude recessive inheritance so far, since the importance of penetrance is unknown and a simple gene model is improbable, c) yielded also negative lod scores (which did not reach the elimination level of -2) for an intermediary genetic model.

[Blood group expertises including related presumptive fathers (author's transl)]. / Blutgruppengutachten mit nahe verwandten Eventualvätern.

In four affiliation cases with two or three brothers involved as putative fathers, exclusions of non-fathers were obtained by the application of the so-called isoelectrofocusing systems (twice by PiM subtypes, once by PGM1, TfC and Gc1 subtypes) as well as by GPT and the extended Gm system. In three cases a discrimination between the involved brothers was possible, in a fourth case both brothers were excluded from paternity. Special problems of anthropological expertises and of biostatistics in cases with closely related putative fathers are discussed. Possibilities to refute false statements of the alleged men, claiming sexual intercourse of the mother with their own relatives in order to invalidate positive biostatistical evidence, are pointed out.

[HLA typing of donor corneas with extended post mortem time]. / HLA-Typisierung von Spenderhornhäuten mit hohen post-mortem-Zeiten.

In case of high-risk patients cornea transplantation should be carried out using HLA-matched donor corneas to minimize the risk of rejection. HLA typing using blood lymphocytes of the donor is impossible because of the long post-mortem times. Alternatively, HLA typing can be performed using retinal pigment epithelium (RPE). Nevertheless, this method is limited by increasing post-mortem times. The aim of the present study was the optimization of culture conditions for RPE cells isolated from donor eyes with long post-mortem times. The HLA type should be evaluated during an acceptable period of organ preservation of the corresponding cornea. In different steps the method for isolation of the cells was optimized and a growth medium for RPE cells was established. Various supplements, including uvea-conditioned medium, were assessed using growth assays. The optimization of culture conditions led to an increase in the estimation of the complete HLA I and HLA II antigens from 36% to 74%. The time needed for the typing-procedure could be reduced from 38 to 17 days (average). At present 30% of the donor tissue with long post-mortem times can be typed in less than 14 days.

[Risk of infection caused by homologous blood transfusion]. / Infektionsrisiko durch Fremdblutspende.

Compared with other nosocomial infections, transfusion-associated infectious risks are of minor relevance in the Federal Republic of Germany today. The actual spectrum of measures for an optimal degree of safety of blood components comprises a careful donor selection, highly sensitive and specific screening assays for viral parameters, inactivation and quarantine procedures for plasmatic preparations. Further possible improvements, both in the blood bank and clinical setting are discussed with regard to the inevitable residual risks of hemotherapy.

Comparison of flow cytometry vs. a haematology cell analyser-based method to guide the optimal time-point for peripheral blood stem cell apheresis.

BACKGROUND AND OBJECTIVES: For timing the onset of apheresis, parameters obtained by flow cytometry and by a haematological cell analyser were compared. MATERIALS AND METHODS: Haematopoietic cell counts (n = 159) were performed by two different methods; CD34 analyses by flow cytometry, immature myeloid information (IMI) and human progenitor cell counts (HPC) by a haematological cell analyser. RESULTS: Comparing the IMI total results with CD34+ analyses (n = 159) revealed a correlation of r = 0.46 (P < 0.05). Similar results were obtained for HPC (r = 0.44; P < 0.05). CONCLUSION: The haematology analyser-based method does not allow the precise determination of absolute haematopoietic stem cell numbers and is thus not able to replace flow cytometry for the monitoring of peripheral blood stem cell counts.

[Blood group expert evaluation: relation between the extent of testing and the reliability of paternity determination. Reflections on revision and guidelines]. / Das Blutgruppengutachten: Beziehungen zwischen Untersuchungsumfang und Zuverlässigkeit der Vaterschaftsfeststellung. Uberlegungen zur Neufassung der Richtlinien.

In affiliation cases a combined exclusion chance for non-fathers of 99.995% is obtained by the examination of well-established blood group systems. In complicated cases, i.e. if known putative fathers are unavailable, the biostatistical limits for the ascertainment of paternity are obviously very high. They have to be determined by court in each particular case. For routine cases the application of an extended basic blood group expertise, including 19 systems with an combined exclusion chance of 95.17% is considered to be sufficient. The analysis of 263 own filiation cases from 1979 to 1982 yielded an average realistic prior probability of paternity of 83.3%, in 52 many-man affairs even of 90.3%. A similar percentage (89.5%) was observed in 67 two-man affairs of contested legitimacy. Since the father is rarely found among men included at a later stage the rate of children without known father is estimated at 5-15%.

PiT: a new allele in the alpha 1-antitrypsin system.

In a genetic investigation of the population in Hessen, Germany, we found a family with a new, rare allele in the Pi system (alpha 1-antitrypsin). According to electrophoretic analysis and isoelectric focusing patterns, it is designated PiT. A pedigree study suggests autosomal codominant inheritance. The serum concentration of six heterozygous carriers of this allele (phenotype M1T or M2T) revealed normal alpha 1-antitrypsin levels.

A third common allele in the transferrin system, TfC3, detected by isoelectric focusing.

Further genetic heterogeneity of the transferrin system (Tf) was revealed by prolonged isoelectric focusing (PAGIF) of human sera on polyacrylamide gels (pH 4--6.5). One of the two common subtypes of TfC, designated previously as TfC1, is split into TfC1 and the new subtype, TfC3. The gene product of TfC3 has an isoelectric point between C1 and C2. In our sample (n = 252) seven phenotypes, TfC1, C2-1, C2, C3-1, C3-2, C3, and C1B2, were observed and the following frequencies calculated: TfC1 = 0.795, TfC2 = 0.155, TfC3 = 0.042, and TfB2 = 0.008. Family studies (n = 25) indicate an autosomal codominant mode of inheritance. The six TfC phenotypes are present after treatment of sera with neuraminidase and can be revealed in part by PAGIF and subsequent immunofixation.

The efficacy of modern blood group genetics with regard to a case of probable superfecundation.

The reported case of probable superfecundation is apparently the most extensively tested of all published cases: More than 40 specificities of the HLA Loci A and B, 5 of the Locus C, Bf and GLO I, furthermore ABO, MNSs, DCcCwEe, K 1--6, Fya, Fyb, Jka, Jk5, Lu3, Hp, Gc, Gm 1, 2, 4, 5, 21, Km 1, C3, Tf including C-subtypes, Pi, Bg, ACP, PGM1 including the extended polymorphism by PAGIF, AK1, ADA, 6-PGD, EsD, GPT, GALT, CAII and Hb. A reciprocal exclusion for putative father 1 and putative father 2 to the twin 2 and the twin 1, respectively, has been reached in 7 blood group systems: HLA, MNSs, Lu, Hp, Tf, Bg and GPT. The random probability for the fatherhood of an unknown third man suitable for both twins is extremely low among whites (1 : 10(8)) or blacks (1 : 10(9)). The development of paternity investigations within the last two years, mainly basing on isoelectrofocusing techniques, is shortly reported. The following systems are preferably involved: PGM1 (4 common alleles), Gc (including 1F and 1S), Pi (3 common subspecificities of M), C6, Tf (3 common subspecificities of C), FUC, Apo E, C2.

[Initial experiences with HCV detection and confirmation tests of the 3rd generation]. / Erste Erfahrungen mit Anti-HCV-Such- und Bestätigungstests der 3. Generation.

We compared the reactivity of a new 3rd-generation anti-HCV enzyme immunoassay (EIA) using an additional, nonstructural antigen from the NS5 region, with a currently used 2nd-generation anti-HCV EIA, by investigating 419 serum specimens from healthy blood donors with normal liver function and 256 samples from patients at risk for hepatitis C infection; 2 of 419 blood donors and 1 of 256 patients were reactive to the NS5 region antigens only (later confirmed by Western blot and RP-2 RIBA), whereas 1 patient was reactive with the 2nd-generation EIA only (antibodies against c33c). We further evaluated the recently developed anti-HCV RP-2 RIBA by testing samples, that were indeterminate by the 2nd-generation RIBA. Nine of 15 patients were identified as reactive by RP-2 RIBA, 6 remained indeterminate. Of the 6 blood donor samples tested, 2 were found reactive, 1 remained indeterminate, and 3 were negative. These test results suggest a higher sensitivity and specificity of the RP-2 RIBA, whereas the relevance of an isolated NS5 EIA reactivity remains to be established.

Properdin factor b-polymorphism in the population of Hessen, Germany.

The polymorphism of properdin factor B (Bf, C3 proactivator) in a population sample from Hessen, Germany has been investigated by agarose gel electrophoresis and immunofixation. In 522 unrelated individuals seven different phenotypes were observed and the following allele frequencies calculated: BfS = 0.7998, BfF = 0.1772, BfS0.7 = 0.0163 and BfF1 = 0.0077. Investigations of 100 families with 198 children and 30 mother-child combinations support the assumed autosomal codominant way of inheritance.