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Biomedical subjects

M A Persson

Publications and source records attributed to M A Persson.

At least 19 recordsLinked to original sources
PubMed1997

Patients infected with the same hepatitis C virus strain display different kinetics of the isolate-specific antibody response.

The antibody response to the hypervariable region of the E2 protein (HVR1) of hepatitis C virus (HCV) was studied in 5 patients who were infected by a common virus strain during an outbreak in a hemodialysis unit. Two patients resolved the infection, while 3 developed chronic HCV infection. For studying the antibody response to HVR1 during the early phase of infection, a Western blot assay using recombinant phage displaying HVR1 was developed. The 2 patients with resolving infection had a more rapid antibody response to HVR1 than did the patients developing chronic infection. Anti-HVR1 antibodies were repeatedly absent in 1 of the chronically infected patients. Antibodies to recombinant E2 protein occurred later than the anti-HVR1 antibodies and did not correlate with resolution of the infection. Thus, the present results suggest that early appearance of antibodies to the HVR1 may predict clearance of HCV infection.

Amino Acid Sequence↗
PubMed1996

Human antibodies from phage libraries: neutralizing activity against human immunodeficiency virus type 1 equally improved after expression as Fab and IgG in mammalian cells.

Human antibodies against HIV-1 have been sought to study neutralization events on the molecular level, and for possible use in passive immune intervention. The development of phage display techniques has opened the possibility of rapidly generating human monoclonal antibodies with desired specificities. We and others have isolated human HIV-1 neutralizing antibody fragments using this technique. Bacterial expression of isolated clones does, however, differ broadly both in expression levels and functional activity. In addition, intact IgG cannot be expressed in bacteria. By transferring the genes of isolated Fab clones to a mammalian expression system we could perform a comparison of functional activity between Fab expressed in bacterial and mammalian cells, as well as Fab and whole IgG. Fab fragments expressed in mammalian cells showed increased virus neutralizing activity compared to the same Fab clones expressed in Escherichia coli, underlining the inefficiency of procaryotic expression. No difference in HIV-1 neutralizing capacity was detected between monovalent (Fab) and divalent (whole antibody) reagents expressed in CHO cells. Thus, bivalency does not always confer improved neutralization efficacy.

Animals↗
PubMed1996

Peptides isolated from random peptide libraries on phage elicit a neutralizing anti-HIV-1 response: analysis of immunological mimicry.

Peptides binding to a murine, human immunodeficiency virus type 1 (HIV-1) neutralizing monoclonal antibody (F58/H3) were isolated from two random peptide libraries expressed on the surface of phage. The antibody was originally elicited by immunization with HIV-1 envelope protein gp120LAI, and has previously been shown to interact with the -I-GPGRA- motif of the V3 loop. The peptide libraries consisted of nine or 15 random amino acid residues flanked by two cysteines, and fused to the amino terminal end of the cpIII protein on the filamentous phage. Selection of specific peptides was carried out in three rounds, with decreasing antibody concentration. An expected peptide motif -GPGRA-, a similar segment, -GPAR-, and two unrelated motifs -FRLLG- and -WRM/ALG- were selected. Binding of antibody was tested both to synthetic peptides in solution, and the corresponding peptide on phage. The GPXR motifs bound in both formats, while the FRLLG bound antibody only when present on the phage The reactivity of peptides on phage was highly dependent on an intact disulphide bond between the cysteines flanking the peptide. The molecular mimicry of the found motifs was tested by immunizing mice and rabbits with conjugated synthetic peptides or peptide on phage. In mice, peptide-specific antisera were raised, but no reactivity to the whole protein (gp120) was detected. In rabbits, however, this was accomplished with the -GPGRA- containing peptide when present on phage. In addition, this antisera precipitated virus particles, and neutralized HIV-1SF2 virus in vitro.

Animals↗
PubMed1995

Frequent patient-to-patient transmission of hepatitis C virus in a haematology ward.

Blood transfusion is a well-documented route of transmission of hepatitis C virus (HCV). However, a persisting high frequency of HCV infections was recorded in our haematology ward even after screening of blood donors had been introduced. We investigated the viral strains in 37 patients with haematological malignant diseases who had developed hepatitis C when treated in the ward during 1990-93. 17 of the patients acquired hepatitis C despite being transfused only with blood components screened by second-generation anti-HCV tests. The viral strains were characterised by PCR genotyping and nucleotide sequencing of the hypervariable region of the E2 gene. Five clusters of closely related or identical viruses were found involving 2, 3, 4, 6, and 15 patients, respectively. Blood components could be ruled out as the common source of infection because no donor had given blood to all patients sharing a specific strain, and even donors whose blood had been given to several patients were negative for HCV RNA. All patients in each cluster had been treated in the ward during overlapping periods. These findings suggest that despite strict hygienic control, HCV transmission occurred between patients treated in the same hospital setting, as has previously been reported in a smaller group of haemodialysis patients.

Adult↗
PubMed1995

Chimeric macaque/human Fab molecules neutralize simian immunodeficiency virus.

A collection of simian immunodeficiency virus (SIV) neutralizing recombinant Fab fragments was generated using the combinatorial antibody library approach. Functional antibody fragments efficiently expressed in Escherichia coli were identified only in the form of chimeric macaque heavy chain gamma 1 and human light chain kappa. The gamma 1 and kappa chains were derived from a clinically healthy long-term surviving SIVsm-infected cynomolgus macaque and from an asymptomatic HIV-2 seropositive individual, respectively. The combinatorial library was constructed on the surface of filamentous phage using the pComb3 phagemid vector and screened against purified SIVsm surface glycoprotein (gp148). Twelve chimeric clones reacting with the antigen were isolated. Six of these clones showed a pronounced neutralizing activity against SIVsm with effects at concentrations of 0.01-0.1 micrograms/ml. All neutralizing Fab fragments were clonally unrelated as demonstrated by nucleic acid sequencing. These potent neutralizing reagents will be used for prophylactic and therapeutic immune intervention of lentivirus infection in macaques and to map neutralizing determinants of SIV.

Amino Acid Sequence↗
PubMed1994

Hepatitis C transmission in a hemodialysis unit: molecular evidence for spread of virus among patients not sharing equipment.

Three cases of simultaneous seroconversion to hepatitis C virus (HCV) in a hemodialysis unit initiated the investigation of the viral strains of 14 seropositive patients in the unit by nucleotide sequencing. The results showed that five patients had been infected with the same viral strain, and indicated that two other patients were sharing a second strain. Transmission was not related to blood transfusions and not associated with the dialysis machines, but occurred between patients treated on the same shift. The number of cases was higher than expected from the serological data. Thus, spread of virus may occur at high frequencies in environments where parenteral routes are made accessible, in spite of rigorous preventive measures. This may raise concern that non-transfusion associated spread of HCV may be present and unnoticed in several hospital settings.

Base Sequence↗
PubMed1993

Combinatorial libraries.

Combinatorial antibody libraries, in which PCR amplified immunoglobulin light and heavy chain DNA are randomly recombined irrespective of their pairing in vivo into a vector and subsequently expressed in E. coli, have quickly become a very productive tool to generate monoclonal antibodies from various species. It has been drastically improved by utilizing phage display technologies in the selection process of specific antibodies. A brief summary of current techniques, critical published experiments showing the versatility of these systems with emphasis on human antibodies and discussions on chain preference, affinity maturation and the advent of semisynthetic and non-immune libraries will be presented.

Animals↗
PubMed1992

[A new genetic technology makes the production of human monoclonal antibodies simpler].

New advances in DNA technology have enabled repertoires of antibodies to be cloned and expressed in bacteria (E coli), thus creating libraries of antibody DNA. In principle, heavy and light chain DNA is amplified from lymphocyte RNA and combined into vectors that enable antibody molecules to be expressed on the surface of phage particles which carry the DNA for the corresponding antibody in their genomes. The system constitutes an ideal tool for use in selecting even rare antibodies from libraries of millions. The new advances have greatly facilitated the generation of monoclonal antibodies against various substances. In particular, human monoclonal antibodies are now more easily obtained with DNA technology than with the methods previously used. Many of the antibodies generated with the new techniques were derived from lymphocyte RNA from immune donors, though the isolation of specific antibodies from libraries originating from donors who have never been in contact with the particular antigens has recently been reported. Moreover, the binding characteristics of the immunoglobulins isolated have subsequently been enhanced in vitro. Thus, modern antibody cloning techniques give promise of allowing the generation of a wide range of specific antibodies from a single, diverse library of antibodies kept 'on the shelf' in the laboratory, without needing to immunise humans or animals. The impact on biomedical science is likely to be profound, as specific reagents for use in research or therapy will be easily obtained, irrespective of whether human or other mammalian antibodies are required.

Antibodies, Monoclonal↗
PubMed1992

Recombinant human Fab fragments neutralize human type 1 immunodeficiency virus in vitro.

A panel of 20 recombinant Fab fragments reactive with the surface glycoprotein gp120 of human type 1 immunodeficiency virus (HIV-1) were examined for their ability to neutralize MN and IIIB strains of the virus. Neutralization was determined as the ability of the Fab fragments to inhibit infection as measured in both a p24 ELISA and a syncytium-formation assay. One group of closely sequence-related Fab fragments was found to neutralize virus in both assays with a 50% neutralization titer at approximately 1 micrograms/ml. Another Fab neutralized in the p24 ELISA but not in the syncytium assay. The other Fab fragments showed weak or no neutralizing ability. The results imply that virion aggregation or crosslinking of gp120 molecules on the virion surface is not an absolute requirement for HIV-1 neutralization. Further, all of the Fab fragments were shown to be competitive with soluble CD4 for binding to gp120 and yet few neutralized the virus effectively, implying that the mechanism of neutralization in this case may not involve receptor blocking. The observation of a preponderance of high-affinity Fab fragments with poor or no neutralizing ability could have implications for vaccine strategies.

Amino Acid Sequence↗
PubMed1991

A large array of human monoclonal antibodies to type 1 human immunodeficiency virus from combinatorial libraries of asymptomatic seropositive individuals.

A panel of human monoclonal antibody Fab fragments has been generated against the surface glycoprotein gp120 of type 1 human immunodeficiency virus (HIV) by antigen selection from a random combinatorial library expressed on the surface of filamentous phage. The library was prepared from 5 ml of bone marrow from an asymptomatic individual who has been HIV-positive for 6 years. The antibodies have high affinity for antigen (mostly with affinity constants of greater than 10(8) M-1) and notable sequence diversity. Given appropriate donor selection, the methods described should allow the generation of antibodies for the evaluation of passive immunization as a therapy for AIDS.

Amino Acid Sequence↗
PubMed1991

Expression of a human monoclonal anti-(rhesus D) Fab fragment in Escherichia coli with the use of bacteriophage lambda vectors.

A human anti-(rhesus D) antibody (IgG1 lambda) Fab fragment was cloned from an Epstein-Barr-virus-transformed cell line and expressed in Escherichia coli with the use of bacteriophage lambda vectors. The cloned protein is active in binding to human erythrocytes and permits the development of a recombinant reagent for the prevention of haemolytic disease of the newborn. The method offers a rapid and effective means of rescuing human Fabs from potentially unstable cell lines secreting human antibodies.

Antibodies, Monoclonal↗
PubMed1991

Generation of diverse high-affinity human monoclonal antibodies by repertoire cloning.

Combinatorial libraries of antibody heavy and light chains derived from the peripheral blood lymphocytes of an individual immunized with tetanus toxoid have been expressed in Escherichia coli by using phage lambda vectors. Screening of the libraries allowed identification of a large number of human monoclonal Fab fragments specific for tetanus toxoid. Initial studies suggested considerable sequence diversity in these antibodies. The method should allow the generation of many human monoclonal antibodies of interest and the dissection of human humoral immune responses.

Amino Acid Sequence↗
PubMed1990

Increased yield of antibody-producing murine spleen cell hybridomas from fusions cultured in medium supplemented with mouse serum.

The number of monoclonal antibody-producing murine hybridomas were increased 7-10-fold in three separate fusions when supplementing the FBS-containing, or serum-free, growth factor-containing, culture medium with 5% normal mouse serum. The beneficial effect was seen in fusions of A/J and 129GIX+ spleen cells, from primary and secondary responses, and when using SP2/0 or P3X63Ag8U.1 as the fusion partner.

Animals↗
PubMed1990

Cerebrospinal fluid interleukin-6 activity in HIV infection and inflammatory and noninflammatory diseases of the nervous system.

Interleukin-6 (IL-6) activity was measured in the cerebrospinal fluid (CSF) of patients at different stages of human immunodeficiency (HIV) virus infection and of patients with multiple sclerosis (MS) or other inflammatory (OID) and noninflammatory neurological diseases (OND). In the advanced stages of HIV infection and in OID, IL-6 was detected more frequently (80 and 75% of the cases) and at higher concentrations than in the early stages of HIV infection. MS and OND (44, 48, and 44% of cases). Analysis of CSF and paired sera indicated that IL-6 production can be compartmentalized to either of the fluids. Evidence that altered blood-brain barrier functions can, at least in part, influence the CSF IL-6 levels was found in OID patients. No association was evident between intrathecal immunoglobulin synthesis and CSF IL-6 levels. Interleukin-1 (IL-1) levels were detectable in a minority of the samples from neurological patients; one OID patient had high levels of both CSF IL-1 and IL-6.

Cerebral Infarction↗
PubMed1990

The neuropeptide substance P stimulates production of interleukin 1 in human blood monocytes: activated cells are preferentially influenced by the neuropeptide.

Substance P (SP) has recently been reported to induce interleukin 1 (IL-1) production by human monocytes. This was confirmed in our experiments with human monocytes cultivated in the presence of SP or SP together with lipopolysaccharide (LPS). In addition, a wide variability of cell response to the neuropeptide was noticed. Three out of twelve cell cultures were directly stimulated by SP to release IL-1, while four additional cultures needed prestimulation with suboptimal doses of LPS, and no effect was seen in the five remaining experiments. The data may suggest that preferentially activated monocytes respond to SP. The production of IL-1 by SP-stimulated monocytes is of great interest considering the broad spectrum of activity of IL-1 and the increasing evidence of sensory neuron involvement in acute and chronic inflammatory responses.

Cells, Cultured↗