Reiner Thomssen - Academia.edu (original) (raw)
Papers by Reiner Thomssen
Med Microbiol Immunol, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Med Microbiol Immunol, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Acta Paediat, 2008
27 consecutive cases with acute peripheral facial palsy were studied for Lyme borreliosis. In 16 ... more 27 consecutive cases with acute peripheral facial palsy were studied for Lyme borreliosis. In 16 out of 27 children Lyme borreliosis could be diagnosed by detection of specific IgM antibodies in CSF. CSF findings allow a clear distinction according to etiology. All children with facial palsy due to Lyme borreliosis revealed lymphocytic CSF pleocytosis, whereas in cases of unknown etiology CSF was usually normal. Bilateral facial palsy occurred only in children with Lyme borreliosis. All cases with a positive history of tick bite and/or erythema migrans in the head-neck region showed ipsilateral neurological affection suggesting a direct invasion via the affected nerve by Borrelia burgdorferi.
Medical Microbiology and Immunology, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Journal of Virology, Mar 1, 1994
The hepatitis B virion is a spherical double-shelled particle carrying three surface proteins (la... more The hepatitis B virion is a spherical double-shelled particle carrying three surface proteins (large [L], middle [M], and small [S]) in its envelope. All three proteins are translated from a single open reading frame by means of three different in-frame start codons from unspliced mRNAs. This organization defines three protein domains (pre-Sl, pre-S2, and S). All three domains together form the L protein, whereas the M protein consists of domains pre-S2 plus S. The L and S proteins are both necessary for virion production, whereas the M protein is dispensable, suggesting an important function of the pre-Sl domain in virion morphogenesis. To investigate this point, we created a series of N-terminal-truncated L mutants and tested their ability to substitute for the wild-type L protein in virion formation. We found that the constructs fell into two classes.
We generated a large number of mutations in the hepatitis B virus (HBV) core gene inserted into a... more We generated a large number of mutations in the hepatitis B virus (HBV) core gene inserted into a bacterial expression vector. The new mutagenesis procedure generated deletions and insertions (as sequence repeats) of various lengths at random positions between M1 and E145 but not substitutions. The R-rich 30-amino-acid C-terminal domain was not analyzed. A total of 50,000 colonies were tested
Med Microbiol Immunol, 1959
Ergebnisse der Mikrobiologie Immunitätsforschung und Experimentellen Therapie, 1961
DMW - Deutsche Medizinische Wochenschrift, 1961
Scandinavian Journal of Infectious Diseases, 1991
We have developed a modified ELISA for the detection of anti-Borrelia burgdorferi (Bb) antibodies... more We have developed a modified ELISA for the detection of anti-Borrelia burgdorferi (Bb) antibodies based on a peroxidase enzyme labelled antigen (ELAT). Microtiter plates were coated with antigen of Bb. The immunoglobulins of the serum samples were bound to the antigen and specific antibodies were detected by an enzyme labelled antigen. The test principle facilitates the recognition of specific antibodies in different collectives of human and animal sera. We performed epidemiological studies with the ELAT on 231 sera from mothers in maternity wards (9.5% positive), 219 patient sera sent to the Bb routine diagnostics (15% positive) and 230 sera from forestry workers (21.3% positive). We further investigated sera from red deer from South Lower Saxony which remained 55% Bb-antibody positive; deer were 37% and fallow deer were 29% positive.
Medical Microbiology and Immunology, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Medical Microbiology and Immunology, 2002
In most sera of hepatitis C virus (HCV)infected patients b-lipoproteins are bound to HCV RNA-carr... more In most sera of hepatitis C virus (HCV)infected patients b-lipoproteins are bound to HCV RNA-carrying material, most often simultaneously with immunoglobulins (IgG, IgM) and sometimes additionally with high-density lipoproteins, forming complexes of low density (1.04-1.06 g/ml). To separate HCV particles from bound material, we tried to destroy the lipoprotein enzymatically by incubating HCV-positive human sera with lipoprotein lipase derived from Pseudomonas spp. (LPL-Ps). After this treatment, titers of HCV RNA in human sera (determined by a simple semiquantitative reverse transcription-PCR assay) were strongly reduced, regardless of whether primers of the NTR or NS5 region were used. Inactivation of HCV RNA could be inhibited by the addition of RNAguard to the serum-enzyme mixture. The lytic effect of the LPL-Ps preparation could be inhibited by tetrahydrolipstatin. Hence LPL-Ps seems to disrupt the HCV particle structure, including the putative core of the virus, and makes HCV RNA sensitive for RNase present in the reaction mixture. HBV was not destroyed by LPL-Ps. Porcine pancreatic lipase had no effect on HCV. The implications of these observations for the structure and biology of HCV and for its stability and inactivation in human sera are discussed. Keywords Hepatitis C virus AE Lipoproteins (low density, very low density) AE Lipoprotein lipase AE Disruption of HCV particles AE Apolipoprotein B100 Lipases and lipase inhibitor The following lipases were used: lyophilized lipoprotein lipase (LPL-Ps) from Pseudomonas spp.; EC 3.1.1.34; Merck no. 1.05389.0001; 10,000 U/30 mg [stock solution: 8 mg in 400 ll 0.2 M TRIS buffer pH 8.0 (6667.5 U/ml)]; and porcine pancreas lipase 16,000 U/mg; (9001-62-1); lot no. 84764; cat. no. 100 823,
Journal of Virological Methods, 1996
The principle and practice of the polymerase chain reaction (PCR) has had a major impact on medic... more The principle and practice of the polymerase chain reaction (PCR) has had a major impact on medical research. This is a powerful method but it does have its limitations, especially for clinical diagnostic work. We describe some improvements of hepatitis C virus (HCV) amplification such as simplification of specimen preparation, elimination of false negative reactions influenced by point mutations, and fluorimetric detection. The aim of the method is to make the procedure as easy and as inexpensive as possible for routine laboratories and for blood screening. After rapid chemical denaturation of the clinical specimen with guanidine thiocyanate and simultaneous hybridization of biotinylated primers to template HCV RNA, the product was fixed to streptavidin-coated magnetic beads and potential inhibitors were removed in easy washing steps. To eliminate the influence of point mutations within the primer binding sites, primer sets with different lengths at their 3'-end were developed for capture, reverse transcription, and amplification of genomic fragments by PCR. Positive results were identified by fluorescence staining. The low cost of the method allows the quantitation of templates by testing of dilution series as is common in microbiological laboratories.
Journal of Medical Virology, 1993
The prevalence and time course of the occurrence of antibodies to the hepatitis B virus polymeras... more The prevalence and time course of the occurrence of antibodies to the hepatitis B virus polymerase (anti-HBpol) were investigated in acutely and in chronically HBV-infected individuals by using recombinant HBpol protein for Western blot analysis. One group consisted of 19 patients who were acutely infected and recovered completely. Five of these patients (26%, 69 serum samples examined) exhibited anti-HBpol. Among those anti-HBpol positive patients, recovery from the disease was combined with a complete loss of this antibody. In contrast, in a second group of 15 individuals who developed chronic hepatitis B, 13 (87%, 102 serum samples examined) had anti-HBpol during the acute phase of the disease. The difference between the anti-HBpol prevalence rates of the two patient groups is statistically significant (Exact Fisher test, P < .002), implying that the occurrence of anti-HBpol may be indicative of a potential chronic course of hepatitis B. Remarkably, anti-HBpol was found in one case of a clinically suspected hepatitis B in which no other serological HBV parameters were found. This serum sample was positive in HBV PCR, supporting a possible diagnostic value of anti-HBpol. 0 1993 Wiley-Liss, Inc.
Journal of General Virology, 1979
The intracellular state of Marek's disease virus (MDV) DNA was investigated in two permanent chic... more The intracellular state of Marek's disease virus (MDV) DNA was investigated in two permanent chicken cell lines: HPRS-I and MSB-I. The HPRS-I line was established from an ovarian lymphoma of a chicken with Marek's disease and is a virus-non-productive line, while the MSB-I line originates from another animal with a Marek's disease splenic lymphoma and is a low producer line. By repeated isopycnic centrifugation in CsC1, MDV DNA in the HPRS-I line showed properties of integrated DNA, whereas in cells of the productive MSB-I line both integrated and free virus DNA appeared to be present. Under denaturing conditions (o-I M-NaOH) the virus DNA remained associated with the cellular DNA as revealed by equilibrium centrifugation in CsC1 and hybridization of the DNA in each single fraction with 32P-labelled complementary RNA transcribed from the DNA of the GA strain of MDV. Shearing of HPRS-I DNA to a tool. wt. of about 8 x IO ~ released only part of the virus DNA to the density of free virus DNA, while a large proportion of MDV DNA could still be localized at the density of cellular DNA. Sedimentation velocity experiments with HPRS-I and MSB-I DNA originally fractionated on CsC1 gradients revealed integrated virus DNA sequences in both cell lines and an additional peak of virus DNA at the position of free linear MDV DNA in the MSB-I line. No fast-sedimenting virus DNA molecules with properties of covalently closed circular structures could be detected. Further evidence for the presence of integrated virus DNA sequences in MDV-transformed cells was provided by the Hirt (1967) precipitation procedure.
Journal of Experimental Medicine, 1990
In sera from patients with acute EBV, infection and the clinical symptoms of infectious mononucle... more In sera from patients with acute EBV, infection and the clinical symptoms of infectious mononucleosis antibodies of the Ig class M were found that are directed against two cellular proteins. The molecular mass of these proteins was determined to be 29 (p29) and 26 kD (p26), respectively, in SDS-PAGE. P29 was identified as part of the glycolytic enzyme triosephosphate isomerase (TPI) by comparison of the NH2-terminal amino acid sequences. A purified antibody against TPI induces a 51Cr release from human erythrocytes. Possibly, anti-TPI causes hemolysis, which is an infrequent but serious symptom of infectious mononucleosis.
International Journal of Cancer, 1991
Sera from 118 women of 33 to over 90 years of age, with or without a history of cervical squamous... more Sera from 118 women of 33 to over 90 years of age, with or without a history of cervical squamous-cell carcinoma, were examined for the presence of antibodies to HPV-6b, HPV-16 and HPV-18, L1, L2, E4, and E7 gene products by the use of bacterially derived beta-Gal fusion proteins and Western-blot analysis. Among the cervical cancer patients, 29/46 (63.0%) were positive for antibodies to E4 and/or E7 of HPV-16 and/or E7 of HPV-18. In contrast, only 2 of 31 (6.5%) non-genital cancer patients and 4 of 41 (9.8%) healthy individuals were antibody-positive for HPV-16 E4 or E7, while antibodies to the homologous proteins of HPV-18 could not be detected. Prevalence rates of antibodies to the HPV-16/18 late proteins were 25/46 (54.3%) in the cervical carcinoma group, 13/31 (41.9%) among women with non-genital cancer types, and 18/41 (43.9%) among normal, healthy individuals. Antibodies to HPV-6b late gene products ranged between 6.5% and 12.2% in the different patient groups. Antibodies to HPV-6b E4 and E7 were detected only once. By studying an additional control group of 207 women with a different age distribution, age-dependence of antibodies to HPV gene products could be ruled out. Whereas antibodies to late proteins may indicate that, regardless of clinical stage, HPV infections are wide-spread among the female population, the striking difference between the prevalence rates of antibodies to early proteins of HPV-16 and HPV-18 among cervical cancer patients and controls (p less than 0.001) supports the idea of the involvement of these virus types in carcinogenesis of the cervix.
Med Microbiol Immunol, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Med Microbiol Immunol, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Acta Paediat, 2008
27 consecutive cases with acute peripheral facial palsy were studied for Lyme borreliosis. In 16 ... more 27 consecutive cases with acute peripheral facial palsy were studied for Lyme borreliosis. In 16 out of 27 children Lyme borreliosis could be diagnosed by detection of specific IgM antibodies in CSF. CSF findings allow a clear distinction according to etiology. All children with facial palsy due to Lyme borreliosis revealed lymphocytic CSF pleocytosis, whereas in cases of unknown etiology CSF was usually normal. Bilateral facial palsy occurred only in children with Lyme borreliosis. All cases with a positive history of tick bite and/or erythema migrans in the head-neck region showed ipsilateral neurological affection suggesting a direct invasion via the affected nerve by Borrelia burgdorferi.
Medical Microbiology and Immunology, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Journal of Virology, Mar 1, 1994
The hepatitis B virion is a spherical double-shelled particle carrying three surface proteins (la... more The hepatitis B virion is a spherical double-shelled particle carrying three surface proteins (large [L], middle [M], and small [S]) in its envelope. All three proteins are translated from a single open reading frame by means of three different in-frame start codons from unspliced mRNAs. This organization defines three protein domains (pre-Sl, pre-S2, and S). All three domains together form the L protein, whereas the M protein consists of domains pre-S2 plus S. The L and S proteins are both necessary for virion production, whereas the M protein is dispensable, suggesting an important function of the pre-Sl domain in virion morphogenesis. To investigate this point, we created a series of N-terminal-truncated L mutants and tested their ability to substitute for the wild-type L protein in virion formation. We found that the constructs fell into two classes.
We generated a large number of mutations in the hepatitis B virus (HBV) core gene inserted into a... more We generated a large number of mutations in the hepatitis B virus (HBV) core gene inserted into a bacterial expression vector. The new mutagenesis procedure generated deletions and insertions (as sequence repeats) of various lengths at random positions between M1 and E145 but not substitutions. The R-rich 30-amino-acid C-terminal domain was not analyzed. A total of 50,000 colonies were tested
Med Microbiol Immunol, 1959
Ergebnisse der Mikrobiologie Immunitätsforschung und Experimentellen Therapie, 1961
DMW - Deutsche Medizinische Wochenschrift, 1961
Scandinavian Journal of Infectious Diseases, 1991
We have developed a modified ELISA for the detection of anti-Borrelia burgdorferi (Bb) antibodies... more We have developed a modified ELISA for the detection of anti-Borrelia burgdorferi (Bb) antibodies based on a peroxidase enzyme labelled antigen (ELAT). Microtiter plates were coated with antigen of Bb. The immunoglobulins of the serum samples were bound to the antigen and specific antibodies were detected by an enzyme labelled antigen. The test principle facilitates the recognition of specific antibodies in different collectives of human and animal sera. We performed epidemiological studies with the ELAT on 231 sera from mothers in maternity wards (9.5% positive), 219 patient sera sent to the Bb routine diagnostics (15% positive) and 230 sera from forestry workers (21.3% positive). We further investigated sera from red deer from South Lower Saxony which remained 55% Bb-antibody positive; deer were 37% and fallow deer were 29% positive.
Medical Microbiology and Immunology, 1993
Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in h... more Heterogeneities in the density of hepatitis C virus RNA-carrying material (HCV-RNA-CM) found in human sera (1.03-1.20 g/cm3) are attributed to the binding of low-density lipoproteins and/or of IgG. In some sera HCV-RNA-CM seems to be nearly totally bound to beta-lipoproteins and cannot be precipitated by anti-IgG (gamma); in others more than 95% of HCV-RNA-CM is bound to IgG and cannot be precipitated by anti-beta-lipoprotein. Furthermore, there are sera from which HCV-RNA-CM can be completely be precipitated by either anti-beta-lipoprotein or anti-IgG (gamma), pointing to a binding of the two serum proteins to the same HCV-RNA-CM. There are other sera from which HCV-RNA-CM can be partially precipitated by the one or the other antiserum, leaving behind fractions, which are bound to beta-lipoprotein or to IgG. HCV-RNA-CM cannot be precipitated from some sera either by anti-beta-lipoprotein or by anti-IgG (gamma).
Medical Microbiology and Immunology, 2002
In most sera of hepatitis C virus (HCV)infected patients b-lipoproteins are bound to HCV RNA-carr... more In most sera of hepatitis C virus (HCV)infected patients b-lipoproteins are bound to HCV RNA-carrying material, most often simultaneously with immunoglobulins (IgG, IgM) and sometimes additionally with high-density lipoproteins, forming complexes of low density (1.04-1.06 g/ml). To separate HCV particles from bound material, we tried to destroy the lipoprotein enzymatically by incubating HCV-positive human sera with lipoprotein lipase derived from Pseudomonas spp. (LPL-Ps). After this treatment, titers of HCV RNA in human sera (determined by a simple semiquantitative reverse transcription-PCR assay) were strongly reduced, regardless of whether primers of the NTR or NS5 region were used. Inactivation of HCV RNA could be inhibited by the addition of RNAguard to the serum-enzyme mixture. The lytic effect of the LPL-Ps preparation could be inhibited by tetrahydrolipstatin. Hence LPL-Ps seems to disrupt the HCV particle structure, including the putative core of the virus, and makes HCV RNA sensitive for RNase present in the reaction mixture. HBV was not destroyed by LPL-Ps. Porcine pancreatic lipase had no effect on HCV. The implications of these observations for the structure and biology of HCV and for its stability and inactivation in human sera are discussed. Keywords Hepatitis C virus AE Lipoproteins (low density, very low density) AE Lipoprotein lipase AE Disruption of HCV particles AE Apolipoprotein B100 Lipases and lipase inhibitor The following lipases were used: lyophilized lipoprotein lipase (LPL-Ps) from Pseudomonas spp.; EC 3.1.1.34; Merck no. 1.05389.0001; 10,000 U/30 mg [stock solution: 8 mg in 400 ll 0.2 M TRIS buffer pH 8.0 (6667.5 U/ml)]; and porcine pancreas lipase 16,000 U/mg; (9001-62-1); lot no. 84764; cat. no. 100 823,
Journal of Virological Methods, 1996
The principle and practice of the polymerase chain reaction (PCR) has had a major impact on medic... more The principle and practice of the polymerase chain reaction (PCR) has had a major impact on medical research. This is a powerful method but it does have its limitations, especially for clinical diagnostic work. We describe some improvements of hepatitis C virus (HCV) amplification such as simplification of specimen preparation, elimination of false negative reactions influenced by point mutations, and fluorimetric detection. The aim of the method is to make the procedure as easy and as inexpensive as possible for routine laboratories and for blood screening. After rapid chemical denaturation of the clinical specimen with guanidine thiocyanate and simultaneous hybridization of biotinylated primers to template HCV RNA, the product was fixed to streptavidin-coated magnetic beads and potential inhibitors were removed in easy washing steps. To eliminate the influence of point mutations within the primer binding sites, primer sets with different lengths at their 3'-end were developed for capture, reverse transcription, and amplification of genomic fragments by PCR. Positive results were identified by fluorescence staining. The low cost of the method allows the quantitation of templates by testing of dilution series as is common in microbiological laboratories.
Journal of Medical Virology, 1993
The prevalence and time course of the occurrence of antibodies to the hepatitis B virus polymeras... more The prevalence and time course of the occurrence of antibodies to the hepatitis B virus polymerase (anti-HBpol) were investigated in acutely and in chronically HBV-infected individuals by using recombinant HBpol protein for Western blot analysis. One group consisted of 19 patients who were acutely infected and recovered completely. Five of these patients (26%, 69 serum samples examined) exhibited anti-HBpol. Among those anti-HBpol positive patients, recovery from the disease was combined with a complete loss of this antibody. In contrast, in a second group of 15 individuals who developed chronic hepatitis B, 13 (87%, 102 serum samples examined) had anti-HBpol during the acute phase of the disease. The difference between the anti-HBpol prevalence rates of the two patient groups is statistically significant (Exact Fisher test, P < .002), implying that the occurrence of anti-HBpol may be indicative of a potential chronic course of hepatitis B. Remarkably, anti-HBpol was found in one case of a clinically suspected hepatitis B in which no other serological HBV parameters were found. This serum sample was positive in HBV PCR, supporting a possible diagnostic value of anti-HBpol. 0 1993 Wiley-Liss, Inc.
Journal of General Virology, 1979
The intracellular state of Marek's disease virus (MDV) DNA was investigated in two permanent chic... more The intracellular state of Marek's disease virus (MDV) DNA was investigated in two permanent chicken cell lines: HPRS-I and MSB-I. The HPRS-I line was established from an ovarian lymphoma of a chicken with Marek's disease and is a virus-non-productive line, while the MSB-I line originates from another animal with a Marek's disease splenic lymphoma and is a low producer line. By repeated isopycnic centrifugation in CsC1, MDV DNA in the HPRS-I line showed properties of integrated DNA, whereas in cells of the productive MSB-I line both integrated and free virus DNA appeared to be present. Under denaturing conditions (o-I M-NaOH) the virus DNA remained associated with the cellular DNA as revealed by equilibrium centrifugation in CsC1 and hybridization of the DNA in each single fraction with 32P-labelled complementary RNA transcribed from the DNA of the GA strain of MDV. Shearing of HPRS-I DNA to a tool. wt. of about 8 x IO ~ released only part of the virus DNA to the density of free virus DNA, while a large proportion of MDV DNA could still be localized at the density of cellular DNA. Sedimentation velocity experiments with HPRS-I and MSB-I DNA originally fractionated on CsC1 gradients revealed integrated virus DNA sequences in both cell lines and an additional peak of virus DNA at the position of free linear MDV DNA in the MSB-I line. No fast-sedimenting virus DNA molecules with properties of covalently closed circular structures could be detected. Further evidence for the presence of integrated virus DNA sequences in MDV-transformed cells was provided by the Hirt (1967) precipitation procedure.
Journal of Experimental Medicine, 1990
In sera from patients with acute EBV, infection and the clinical symptoms of infectious mononucle... more In sera from patients with acute EBV, infection and the clinical symptoms of infectious mononucleosis antibodies of the Ig class M were found that are directed against two cellular proteins. The molecular mass of these proteins was determined to be 29 (p29) and 26 kD (p26), respectively, in SDS-PAGE. P29 was identified as part of the glycolytic enzyme triosephosphate isomerase (TPI) by comparison of the NH2-terminal amino acid sequences. A purified antibody against TPI induces a 51Cr release from human erythrocytes. Possibly, anti-TPI causes hemolysis, which is an infrequent but serious symptom of infectious mononucleosis.
International Journal of Cancer, 1991
Sera from 118 women of 33 to over 90 years of age, with or without a history of cervical squamous... more Sera from 118 women of 33 to over 90 years of age, with or without a history of cervical squamous-cell carcinoma, were examined for the presence of antibodies to HPV-6b, HPV-16 and HPV-18, L1, L2, E4, and E7 gene products by the use of bacterially derived beta-Gal fusion proteins and Western-blot analysis. Among the cervical cancer patients, 29/46 (63.0%) were positive for antibodies to E4 and/or E7 of HPV-16 and/or E7 of HPV-18. In contrast, only 2 of 31 (6.5%) non-genital cancer patients and 4 of 41 (9.8%) healthy individuals were antibody-positive for HPV-16 E4 or E7, while antibodies to the homologous proteins of HPV-18 could not be detected. Prevalence rates of antibodies to the HPV-16/18 late proteins were 25/46 (54.3%) in the cervical carcinoma group, 13/31 (41.9%) among women with non-genital cancer types, and 18/41 (43.9%) among normal, healthy individuals. Antibodies to HPV-6b late gene products ranged between 6.5% and 12.2% in the different patient groups. Antibodies to HPV-6b E4 and E7 were detected only once. By studying an additional control group of 207 women with a different age distribution, age-dependence of antibodies to HPV gene products could be ruled out. Whereas antibodies to late proteins may indicate that, regardless of clinical stage, HPV infections are wide-spread among the female population, the striking difference between the prevalence rates of antibodies to early proteins of HPV-16 and HPV-18 among cervical cancer patients and controls (p less than 0.001) supports the idea of the involvement of these virus types in carcinogenesis of the cervix.