| 1. |
- Larrick, James W, et al.
(författare)
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Polymemse chain reaction using mixed primers. Cloning of human monoclonal antibody variable region genes from single hybridoma cells
- 1989
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Ingår i: Bio/Technology. - : Springer Science and Business Media LLC. - 0733-222X. ; 7:9, s. 934-938
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Tidskriftsartikel (refereegranskat)abstract
- We describe a general approach to rapidly obtain the DNA sequence encoding the variable region of any immunoglobulin chain using the polymerase chain reaction and a mixture of upstream primers corresponding to the leader sequence, and one downstream primer designed from the conserved nucleotide sequence of the constant region. The approach was applied to five different hybridomas producing human monoclonal antibodies and variable regions for both bold gamma and mu heavy chain and kappa and lambda light chain genes were successfully cloned. cDNA encoding variable regions could be amplified from single hybridoma cells isolated by micromanipulation. This approach will permit analysis of B cell clonal ontogeny, antibody diversity and lymphoma cell progression and heterogeneity. It will also facilitate structural and functional studies of immunoglobulins as well as the rapid construction of chimeric antibodies.
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| 2. |
- Larrick, James W, et al.
(författare)
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Rapid cloning of rearranged immunoglobulin genes from human hybridoma cells using mixed primers and the polymerase chain reaction
- 1989
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Ingår i: Biochemical and Biophysical Research Communications. - 1090-2104. ; 160:3, s. 1250-1256
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Tidskriftsartikel (refereegranskat)abstract
- A general method to directly obtain the DNA sequence of the variable regions of any immunoglobulin chain using a mixture of oligomer primers and the polymerase chain reaction (PCR) is described. Mixed oligonucleotide primers corresponding to the 5′ signal peptide and a conserved 3′ constant region primer were used for enzymatic amplification of each of the heavy and light chain variable regions of a human hybridoma producing a monoclonal antibody recognizing an epitope of gp120 of the human immunodeficiency virus 1. The amplified DNA segments were cloned and the sequence was determined for the heavy chain variable region. This method will greatly facilitate structural and functional studies of immunoglobulins by reducing the effort to clone and sequence the members of the immunoglobulin as well as other multigene families.
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| 3. |
- Ljungqvist, Olle, 1954-, et al.
(författare)
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Evidence of increased gluconeogenesis during hemorrhage in fed and 24-hour food-deprived rats
- 1989
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Ingår i: Journal of Trauma and Acute Care Surgery. - : Lippincott Williams & Wilkins. - 2163-0755 .- 2163-0763. ; 29:1, s. 87-90
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Tidskriftsartikel (refereegranskat)abstract
- Food withdrawal 24 hr before hemorrhage has been shown to increase experimental post-hemorrhage mortality, and survival is associated with the degree of hyperglycemia. Lack of hyperglycemic response has been attributed to depleted glycogen reserves after 24-hr food withdrawal. To investigate the effect of short-term food deprivation on glucose metabolism during hemorrhagic stress, glucose production (rate of appearance, Ra), glucose uptake (rate of disappearance, Rd), glucose clearance, and glucose recycling were investigated in fed and 24-hr food-deprived rats under basal conditions, and during hemorrhagic hypotension using 3-H3-U-C14-glucose. During hemorrhage, blood glucose levels were higher in fed rats. Hemorrhage induced a decrease in glucose clearance irrespective of nutritional state in both 24-hr starved animals and rats in the postprandial state. Calculated glucose recycling increased in both groups after hemorrhage. The results indicate that hemorrhagic stress induces a rapid increase in gluconeogenesis, as reflected by increased glucose recycling.
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| 4. |
- Lopez-Rivas, Abelardo, et al.
(författare)
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Ca2+-mobilizing actions of platelet-derived growth factor differ from those of bombesin and vasopressin in Swiss 3T3 mouse cells
- 1987
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : The National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 84:16, s. 5768-5772
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Tidskriftsartikel (refereegranskat)abstract
- Addition of the mitogenic peptides bombesin and vasopressin to quiescent Swiss 3T3 mouse cells increased the cytosolic Ca2+ concentration without any measurable delay. In contrast, there was a significant lag period (16 +/- 1.2 s) before platelet-derived growth factor (PDGF) increased cytosolic Ca2+ concentration. This lag was not diminished at high concentrations of either porcine or human PDGF. Similar results were obtained in 3T3 cells loaded with quin-2 or fura-2. The differences in the effects of bombesin, vasopressin, and PDGF on Ca2+ movements were also substantiated by measurements of 45Ca2+ efflux and of cellular 45Ca2+ content. Activation of protein kinase C by phorbol esters inhibited Ca2+ mobilization induced by either bombesin or vasopressin. In contrast, phorbol esters had no effect on PDGF-induced cytosolic Ca2+ concentration increase or acceleration of 45Ca2+ efflux. Finally, bombesin and vasopressin caused a rapid increase in the production of inositol 1,4,5-trisphosphate and inositol 1,3,4-trisphosphate, whereas PDGF, even at a saturating concentration, exerted only a small effect. These results indicate that the signal transduction pathways activated by PDGF that lead to Ca2+ mobilization can be distinguished from those utilized by bombesin and vasopressin.
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