Immunoenzyme AB0 blood group determination using a single hair. I

The immunoenzyme technique was used to determine the ABO blood group of strands of human scalp hair. The hair was obtained from 168 individuals of known blood groups (A1: n = 58; A2: n = 11; B: n = 28; O: n = 46; A1B: n = 16; A2B: n = 9). Immunostaining was carried out by using monoclonal anti-A, anti-B and anti-H as primary antibodies. Group-specific staining was clearly observed within the medulla of the hair. The ABO blood group of all hair samples was determined correctly by the Sternberger (PAP) or APAAP (immunoalkaline phosphatase) technique. The present study indicates that immunoenzyme techniques can be regarded as practical methods for determining ABO blood group of hair.     

Determination of hair group by preserved hair cell bulbs

Difficulties appearing in determining hair group were the reason for conducting this research for the purpose of finding additional possibilities to solve the mentioned problem. It was established that the preliminary cell coloring, related with determination of hair sex, does not influence a consequent detection of antigens. Methods or the fixation of material were selected. The most suitable reagents and their titers as well as different time periods of absorption for detecting antigens A and B are offered. All stages of examination are described in detail.     

Determination of ABO(H) blood group substances from finger and toe nails

Thirty-six finger and toe nails were analyzed for ABO(H) blood group substances by the modified absorption elution method. The blood groups from nails were successfully determined in all the samples.     

The ABO blood grouping of a minute hair sample by the immunohistochemical technique

The unlabeled antibody (PAP) immunoperoxidase technique was applied to the ABO blood grouping of human scalp hairs. Hair samples were subjected to longitudinal- or cross-sectioning, thus obtaining suitable samples for subsequent immunostaining. The immunostaining was carried out using rabbit anti-A and anti-B sera as the primary antibodies. With this technique, the group-specific staining which is revealed as a dark brown precipitate was clearly observed within the medullae of the hair shaft, and depending on the presence or absence of these precipitates, respective blood groups of unknown hair samples were determined. At the hair root, on the other hand, positive stainings were observed not only in medullary cells but also in some cortical cells of the keratogenous zone. From the present study, it can be safely said that this technique is of practical use for the ABO blood grouping from a minute (less than 3 mm) hair sample.     

Examinations of ABO bloodgroupings of human hair

In order to bring into play the function of human hair ABO bloodgrouping in the field of medicolegal expertise on material evidence and raise its accuracy, the author, through a test on human hair of 500, makes some emphatic researches on the disposal of hairs, on the application of anti-A and anti-B serum, on the selection of red blood cell indicator and on the elution temperature as well. Five hundred samples of human hair have been examined upon the basis of the improved operation method and through the application of anti-A and anti-B serum, the titer of which being 1:128, and therein, fine results have been obtained.     

Immunochemical studies on blood group A substance from human hair

Blood group A-active substance was extracted from urea-treated human hair uith methanol-ethyl ether 1:1, v/v) or chloroform-methanol (1:1, v/v). The serological activity of blood group A substance in the hair was destroyed by A-decomposing enzyme from Clostridium tertium with concomitant development of blood group H activity. It is concluded therefore that the extract from the hair of group A contained blood group A-active glycolipid with N-acetylgalactosamine as the non-reducing sugar.     

Determination of ABO blood groups by radioimmunoassay using 125I-protein A

Since the crystallizable fragment (Fc) portion of the immunoglobulin G (IgG) molecule is the binding site of Protein A, a radioimmunoassay procedure using 125I-Protein A was developed for identification of the ABO blood groups. The isotope level bound to Group A, B, or AB red cells decreased with the dilution of anti-A or -B, respectively. After sensitization by anti-A plus B in Group O serum, the isotope bindings were observed in Groups A, B, and AB cells, while no significant radioactive count appeared in Group O cells. Furthermore, there was little significant isotope binding in both Group A and B red cells sensitized by the serum from Group A or B blood containing mainly IgM anti-A or -B. A radioimmunoassay using 125I-Protein A is an excellent method for identifying ABO blood groups.     

Immunohistochemical study of the pattern of distribution of ABO blood group substances in male genitalia

The distribution of the ABH antigens was investigated in 11 different sections of the male genitalia in 53 autopsy cases; the peroxidase-antiperoxidase technique was used. Specific staining of the epithelia differed considerably among and between individuals. Nonsecretors showed a tendency toward less staining in the epithelia, whereas in the endothelia there was no difference. A2 cases could be recognized, as the endothelia were marked almost completely with anti-H. In A2B nonsecretor epithelia and endothelia, there was only a minimal reaction with anti-A and anti-H. Spermatozoa were irregularly stained in the ampulla of the vas deferens, whereas in the testis and epididymis no reaction could be found.     

Effect of blood group active micro-organisms on the ABO grouping of human whole saliva

Three saliva samples with false positive ABO grouping results were assayed for blood group active organisms, using a variety of selective media to isolate representative strains from the salivary microflora. Eight out of 40, 8 out of 40 and 4 out of 30 strains from the three samples, respectively, showed blood group activity, which correlated well with the false positive specificities of the saliva samples. In all cases the false reaction only lasted a few days. Investigation of one of these samples before and after the appearance of the false positive activity yielded only one out of 40 blood group active organisms, using the same methods. Similar investigation of two "normal" saliva samples found none out of 40 and one out of 40 blood group active organisms, respectively. It is concluded that occasional false positive ABO grouping reactions of saliva samples are probably caused by the presence of unusually high numbers of blood group active micro-organisms, due to disturbances in the ecological balance of the salivary microflora.     

Comparison of ethyl glucuronide in hair with phosphatidylethanol in whole blood as post-mortem markers of alcohol abuse

Ethyl glucuronide (EtG) is a direct metabolite of ethanol and has been used as a marker of alcohol abuse in both urine and hair. This study investigated the value of EtG testing in post-mortem hair for diagnostic improvement of alcohol abuse in forensic medicine. Material from 70 consecutive medico-legal autopsies was collected in accordance with the recommendations on ethics by the Swedish National Board of Forensic Medicine. A method for determination of EtG in hair samples was developed using ultra performance liquid chromatography/electrospray tandem mass spectrometry (UPLC/ESI-MS/MS; LOQ, 2.5 pg/mg). The result of the EtG analysis was compared with the findings of phosphatidylethanol (PEth) in femoral whole blood, as measured by high performance liquid chromatography with an evaporative light-scattering detector (HPLC-ELSD; LOQ, 0.22 micromol/l). Evaluation of liver histology and anamnestic evidence of alcohol abuse of the deceased were taken in consideration for the interpretation. Measurable levels of EtG were present in 49 of the 70 autopsy cases whereas PEth was present in 36. Thirty-nine cases had EtG levels above the cutoff limit (> or = 30 pg/mg) compared with 29 for PEth (> or = 0.7 micromol/l). Fifteen cases had EtG as exclusive indicator for alcohol abuse compared with four cases for PEth. These findings suggest that measurements of EtG in hair may provide improved diagnostic information on alcohol abuse, due to a long retrospective time-window for detection and stability of EtG in hair in the decaying cadaver. However, an EtG level below the cutoff does not completely exclude previous alcohol abuse.     

Determination of MN blood group from blood stains by electrophoresis and immunoblotting

The determination of blood groups from blood stains is extremely important in medicolegal practice, but there is the possibility of an error in the determination of MN phenotypes by the absorption-elution test. We investigated a new method applying electrophoresis and immunoblotting. As a consequence of various experiments, the most appropriate pretreatment of blood stains was as follows. Blood stains were immersed in physiological saline for 0.5 to 1 h and centrifuged. The supernatant was discarded. The sediment was dissolved in sample buffer (TRIS-buffered physiological saline containing 2% sodium dodecyl sulfate) and followed by thermodegradation. It was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). After transfer to a nitrocellulose membrane by Western blotting, MN phenotypes could be determined accurately from blood stains by an enzyme immunoassay (EIA) using commercially available polyclonal anti-M and anti-N sera. For blood stains more than 1 month old it was not easy to determine the MN phenotypes.