Improved phenotyping of alpha 1-antichymotrypsin (ACT) by isoelectric focusing and immunoprinting: first demonstration of a deficient protein variant in the ACT system.

Genetic variation of human alpha 1-antichymotrypsin (ACT) was investigated in sera using thin-layer polyacrylamide gel isoelectric focusing (pH range 4.0-6.5) followed by immunoprinting with a monospecific anti-human ACT antibody. Sialidase-treated samples showed a microheterogeneous banding pattern which consisted of two major and several additional minor components with isoelectric points between pH 5.0 and 5.3. A population study of 200 unrelated individuals from southern Germany revealed no genetic variation. In a clinical investigation, however, we found a unique banding pattern in a female patient suffering from chronic obstructive pulmonary disease. In comparison with the monomorphic normal type the detected variant phenotype shows two additional bands that have lower intensities and are located cathodically to their major bands. Inheritance of the deficient IEF variant "ACT Bochum" was confirmed by a family study. To our knowledge this is the first genetic ACT mutant to be observed at the protein level.

DR2-positive monozygotic twins discordant for narcolepsy.

Narcolepsy runs in families, and recent research has revealed the human leukocyte antigen (HLA) DR2 to be a genetic marker closely associated with the disease. But, as indicated by family studies, other factors contribute to the pathogenesis of narcolepsy. The investigation of monozygotic twins is the most specific research tool for distinguishing between a multigenetic and a multifactorial pathogenetic model. We present clinical and sleep polygraphic data from two pairs of monozygotic twins, and in addition, from some of their first-degree relatives. In both pairs only one twin suffered from the clinical symptoms of narcolepsy/cataplexy. Only in these subjects did night sleep recordings and a multiple sleep latency test reveal both multiple sleep onset rapid-eye-movement periods (SOREMPs) and short mean sleep onset latencies. However, in two of the asymptomatic, HLA DR2+ relatives, short mean sleep onset latencies during the multiple sleep latency test (MSLT) were observed, and one, HLA DR2- relative showed REM sleep two times during the MSLT. Our results strongly favor a multifactorial pathogenetic model for narcolepsy.

Further evidence of a silent plasminogen (PLG) allele in two paternity cases.

Routine paternity testing has yielded two different cases of an apparent inverse homozygosity in the plasminogen (PLG) system. In one case, the child presented the phenotype PLG A and his putative father the type PLG B. The alleged father could not be excluded from the paternity in 25 additional blood group marker systems (biostatistical probability of paternity W greater than 99.75%). In the other case an incompatibility was found in a mother- child pair. Analysis of PLG was carried out by isoelectric focusing on neuraminidase-treated sera. In both cases the immunologic and functional detection showed weaker banding pattern of the affected PLG types. The assumption of a silent allele in the PLG system was confirmed by quantitative investigations. The allele frequency of PLG*Q0 in the South German population was estimated to be 0.0013. In the same sample the variant PLG A3 has been shown to be polymorphic.

Three new orosomucoid (ORM) variants revealed by isoelectric focusing and print immunofixation.

Phenotypes of orosomucoid (ORM) in human sera have been analysed by isoelectric focusing and print immunofixation. After neuraminidase treatment the band patterns indicated that the polymorphism of the structural locus ORM1 is controlled by three autosomal codominant alleles. According to the previous nomenclature they were called ORM1F1, ORM1F2, and ORM1S. In a study of 272 unrelated individuals from southern Germany, five of the six expected common ORM1 subtypes were observed. Furthermore, we found three ORM variant phenotypes which have not been reported previously. These variants were characterized by additional bands in a cathodal position. One variant had additional double bands and presumably represents a rare ORM1 variant named ORM1S1. Two variants had additional single bands. They were assigned tentatively to the ORM2 gene locus. While the common gene product of ORM2 may be called ORM2A, the two variants are named ORM2B1 and ORM2B2, respectively. ORM2B1 has, thus far, been found only in a single individual; the variants ORM1S1 and ORM2B2 were found in a father-child pair and a mother-child pair, respectively. The frequency for variants tentatively assigned to the ORM2 locus is very low and was calculated to be 0.0037.

Plasminogen (PLG): a useful genetic marker for paternity examinations.

The genetically determined polymorphism of plasminogen (PLG) was analyzed by isoelectric focusing on polyacrylamide gels. For analysis neuraminidase-pretreated sera were used. PLG was developed functionally by activation with urokinase and subsequent lysis of casein in an agar overlay. In a random sample of 957 unrelated healthy individuals from Southern Germany, three common phenotypes, PLG1, 2-1, and 2, and five rare variants were found. The allele frequencies were: PLG*1 = 0.7174, PLG*2 = 0.2780, and PLG*Var = 0.0046. The theoretical exclusion rate in cases of disputed paternity is 16.5%.

Alpha-1-antitrypsin: evidence for a fifth PI M subtype and a new deficiency allele PI*Z augsburg.

The phenotypes of the protease inhibitor (PI) alpha-1-antitrypsin have been analyzed by isoelectric focusing on polyacrylamide gels. With improved resolution by a modified procedure it was possible to demonstrate a fifth PI*M suballele. The bands of PI M5 are located between PI M1 and PI M3. In addition, a further deficiency allele similar to PI*Z was found in a female patient with obstructive pulmonary disease. This variant was provisionally named PI Z augsburg (PI Z aug). Family data confirm a simple codominant mode of inheritance for PI Z aug.

Transferrin subtypes and variants in Germany; further evidence for a Tf null allele.

Isoelectric focusing (IEF) with carrier ampholytes was used for the determination of transferrin C subtypes and transferrin B and D variants in a sample of 1125 unrelated individuals from Southern Germany. The observed TfC allele frequencies were Tf*C1 = 0.7872, Tf*C2 = 0.1365, and Tf*C3 = 0.0675. The rare C subtype C6 was observed twice. A new C subtype, called C10, was observed and identified by IEF with immobilized pH gradients. The rare C subtypes C4 and C8 were also studied by this method. TfB and TfD variants were found with a heterozygous frequency of 1.53%. One new TfD was found which is located between D1 and D2 and therefore named D1-2. Evidence for a Tf null allele was obtained in a child and the putative father; they were considered to be heterozygous for an allele Tf0. The theoretical exclusion rate for paternity examinations was calculated for the Tf system and found to be 17.95%.

Two new Bf S subtypes revealed by isoelectric focusing and immunofixation.

The genetic types of the properdin factor B were analyzed by isoelectric focusing on polyacrylamide gels and subsequent immunofixation. Sera from 516 unrelated, healthy individuals from Southern Germany were examined. Two new subtypes of the Bf*S allele were observed. They were provisionally named Bf*Sb1 and Bf*Sb2(b = basic), since the position of their bands is located slightly towards the cathode. Whereas Bf Sb2 has, thus far, been found only in a single individual, Bf Sb1 was found in five unrelated persons and in a mother and her child indicating a simple codominant mode of inheritance. The combined frequency of the Bf*Sb alleles was calculated to be 0.0067.

PGM1 subtypes determined by agarose gel isoelectrofocusing.

PGM1 subtypes were determined in red cell hemolysates by isoelectric focusing on agarose gel plates. By this modified procedure PGM1 subtypes may be readily classified. Nine of the 10 expected phenotypes were found in a sample of 470 unrelated individuals from Southern Germany. The frequencies for the four alleles were found to be: PGM1(1+) = 0.212, PGM1(1-) = 0.1224, PGM1(2+) = 0.2043, PGM1(2-) = 0.0521.

Pi subtyping by isoelectric focusing: further genetic studies and application to paternity examinations.

Genetic variation of the protease inhibitor (Pi) alpha 1-antitrypsin was analyzed by isoelectric focusing on polyacrylamide gels in a sample of 347 unrelated individuals from Southern Germany. Six common subtypes of PiM were observed as well as the relatively frequent variants PiS and PiZ and the rare variants PiT, Pi less than L, PiL, PiI and PiF. Also, a variant called PiZ1 was found. The frequency of alleles in this sample was PiM1 = 0.6917, PiM2 - 0.1686, PiM3 = 0.0865, PiS = 0.0230, PiZ = 0.0187, and Pi* = 0.0115. In 82 families the distribution of Pi types was in agreement with an autosomal codominant mode of inheritance. The application of Pi classification in cases of disputed paternity is discussed.

Classification of transferrin (Tf) subtypes by isoelectric focusing.

A sample of 450 sera from unrelated individuals from Southern Germany was examined by isoelectric focusing on polyacrylamide gels. Three common subtypes, TfC1, C2-1, and C2, were differentiated. In addition, the rare variants TfB1, B1-2, B2, D1, D1-2, D2, and D3 were observed. The frequencies of the Tf alleles in our sample were found to be: TfC1 = 0.8544, TfC2 = 0.1367, TfB1 = 0.0011, TfB1-2 = 0.0022, TfB2 = 0.0045, and TfD1 = 0.0011. Analysis of 73 parents with 73 children did not show deviations from the expected mode of inheritance. Modification of the method by addition of 0.01 M FeCl3 to the sera prior to examination did, however, reveal further variation and permitted the distinction of six subtypes, C1, C2-1, C2, C3, C3-1, and C3-2.

Properdin factor B-polymorphism. An indication for the existence of a Bf O-allele.

The polymorphism of the properdin factor B (Bf, C3-proactivator, GBG = glycin-rich-beta-glycoprotein) has been investigated by high voltage agarose gel immunofixation electrophoresis in 1115 unrelated persons from Southern Germany. Seven phenotypes were observed; the allele frequencies were calculated as BfS = 0.8094, BfF = 0.1790, BfSI = 0.0094, BfFI = 0.0022. A study of 94 parents with 98 children and 420 mother-child combinations showed no deviation from the assumed autosomal codominant mode of inheritance. In one additional family the findings suggested the existence of a silent allele at the Bf-locus.

The medicolegal usefulness of the Kidd system in exclusion of parentage.

The blood samples of 2741 people from Germany were examined. The gene frequencies of Jka was found to be 0.49345 and of Jkb 0.50655. The theoretical probability of exclusion lies at 18,57%, the practical one only at 7.09%, because in many cases, the presumptive father is also the biological father. It is a system of dependable information. The Kidd system can be used as a classical system for exclusion of parentage.

Population genetics of glyoxalase I (E.C.4.4.1.5) in human erythrocytes.

1025 individuals from Southern Germany were examined. The gene frequencies for GLI1 are 0.4235 and for GLI2 0.5765. These frequencies are compared with those of other authors.

Esterase D polymorphism: description of the "new" allele EsD4.

Two "new" phenotypes of the esterase D system, named EsD 4-1 and EsD 4-2, were observed in a father and his daughter, respectively. An additional allele EsD4 is postulated.

Determination of isozymes of phosphoglycerate-mutase (E.C.2.7.5.3) and enolase (E.C.4.2.1.11) in human erythrocytes.

525 human hemolysates were tested for the isozymic patterns of phosphoglycerate mutase and enolase. Genetic models for interpreting the pherograms are suggested.