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E Haber

Publications and source records attributed to E Haber.

At least 217 records · Page 12Linked to original sources
PubMed1982

Technetium-99m labeling of antibodies to cardiac myosin Fab and to human fibrinogen.

We have developed a method of labeling biologically active labile macromolecules, such as human fibrinogen (HF) and anticardiac-myosin Fab (AM-Fab), with Tc-99m at neutral pH. This method uses dithionite reduction of pertechnetate and subsequent labeling, to test the method with acid-labile macromolecules. Complexes of diethylene triamine pentaacetic acid with macromolecules such as human fibrinogen (D-HF) and anticardiac-myosin Fab (D-AM-Fab) were labeled and utilized in in vitro and in vivo studies. In biodistribution studies, the Tc-99m D-HF had a two-component blood clearance (half-times 1 hr and 15 hr) and was 80--88% coagulable. The Tc-99m AM-Fab retained its immunoreactivity as tested by affinity chromatography; also during in vivo localization in experimental myocardial infarction. This labeling technique provides an easy and efficient approach to the Tc-99m labeling of other biologically active and acid-labile macromolecules.

Animals↗
PubMed1981

Quantitative solid-phase Edman degradation for evaluation of extended solid-phase peptide synthesis.

Quantitative solid-phase Edman degradation was used for the amino acid sequence analysis of synthetic peptidyl-resins prepared by the Merrifield solid-phase procedure. A model peptide, Ala-[3H]Pro-Ala-Gly-Phe-Ala-Gly-, was synthesized on a solid support and was sequenced to measure the efficiency of the solid-phase sequencing protocol used. An average of 92% of the first four residues was removed from the peptidyl-resin as indicated by subtractive amino acid analysis. Quantitation of the radioactive proline residue at cycle 2 revealed that it was efficiently recovered both from the acid conversion procedure (99%) and also following high-pressure liquid chromatography of the phenylthiohydantoin (Pth) amino acid (88%). In order to facilitate identification and quantification of the side chain protected Pth amino acids, we prepared these derivatives and characterized them by high-pressure liquid chromatography. Thereafter, by the use of solid-phase Edman degradation as an analytical procedure, the synthesis of residues 2-118 of the heavy-chain variable region (VH) of a homogeneous rabbit antibody was undertaken. At 10-15-residue intervals during the solid-phase synthesis, samples of peptidyl-resin were removed from the synthesis vessel and sequenced. When gross synthetic errors caused by deletion of amino acids residues were detected, the solid-phase synthesis was terminated and restarted by using modified protocols. A 117-residue peptidyl-resin was prepared finally which possessed the desired amino acid sequence as indicated by a series of solid-phase Edman degradation experiments. In the final degradation experiment on the 117-residue peptidyl-resin, a 92% efficiency for the automatic Edman reaction was measured ([3H]Leu, penultimate amino-terminal residue). We have found two advantages for the concurrent use of solid-phase Edman degradation during an extended solid-phase synthesis: (1) on the basis of the level of error due to incomplete incorporation of amino acids, the solid-phase assembly could be terminated in favor of restarting the synthesis, hence avoiding further work on a defective product and (2) direct verification of incorporation of amino acids, which during acid hydrolysis are destroyed (Cys, Trp) or are deamidated (Asn, Gln), is possible by high-pressure liquid chromatography of the corresponding Pth derivatives.

Amino Acid Sequence↗
PubMed1981

Monoclonal antibodies directed against human myoglobin: characterization and application in a bideterminant radioimmunoassay.

Monoclonal hybridoma cell lines secreting antibodies directed against human myoglobin were selected. Two of these cell lines were grown in mouse ascitic fluid resulting in the production of large quantities of antibody. Antimyoglobin antibodies isolated from the ascitic fluids were employed in the development of the sensitive solid-phase, bideterminant radioimmunoassay for human myoglobin that uniquely recognizes two different epitopes on the same molecule.

Animals↗
PubMed1981

Cellular mechanisms of impaired adrenergic responsiveness in neonatal dogs.

The myocardial responsiveness of conscious, instrumental dogs to exogenously administered isoproterenol and norepinephrine was investigated in neonatal, 6-wk-old, and adult animals. Comparable base-line values for peak left ventricular derivative of pressure with respect to time were observed in all age categories. However, when compared with adult responses, the sympathomimetic amine-induced increases in neonatal left ventricular dP/dt were significantly blunted at each concentration of adrenergic agonist examined, whereas the 6-wk-old puppies displayed an intermediate inotropic response. To investigate the cellular mechanisms of this blunted neonatal response, we correlated physiologic and biochemical measurements of the myocardial responses to catecholamines in each age category. When compared with adult myocardial membrane preparations, neonatal cardiac membranes were characterized in vitro by an increased density of beta-adrenergic binding sites, comparable affinity for adrenergic agonists and antagonists, and an enhanced coupling of adenylate cyclase activation to receptor occupancy. Simultaneous changes in either the serum catecholamine concentration or the membrane content of other intrinsic proteins failed to account for the observed neonatal increase in beta-adrenergic receptor density. These findings are most consistent with a compensatory mechanism of the cardiac cell membrane, whereby an inherent depression in the adrenergic responsiveness of the immature myocardium appears to induce the increase in receptor density and activation of adenylate cyclase.

Adenylyl Cyclases↗
PubMed1980

Isolation of an active heavy-chain variable domain from a homogeneous rabbit antibody by cathepsin B digestion of the aminoethylated heavy chain.

Cathepsin B from bovine liver has been used to cleave the heavy chain of partially reduced and aminoethylated rabbit allotype a1 IgG. Three major cleavages have been identified, one of which appears to be at the peptide bond carboxy terminal to the two adjacent (aminoethyl)cysteine residues at positions 133 and 134. The variable domain of the heavy chain (VH) was isolated by gel filtration from both pooled heterogeneous rabbit IgG and a homogenous rabbit antitype III pneumococcal polysaccharide antibody. This VH inhibited the binding of 125I-labeled (allotype a) IgG to anti-a1 allotypic antibodies. The recombinant molecule consisting of VH and light chain from the homogeneous antibody is active in an antigen binding assay.

Animals↗
PubMed1980

Myocardial infarct imaging of antibodies to canine cardiac myosin with indium-111-diethylenetriamine pentaacetic acid.

Antibodies, by virtue of marked selectivity and affinity, may lend themselves to identification of structures of unique antigenic specificity in vivo. In experimental myocardial infarction in dogs, F(ab')2 fragments of antibodies to cardiac myosin that had been labeled with iodine-131 were shown to localize within the lesion. Because the energy characteristics of iodine isotopes are not ideal for imaging with a gamma camera, a new method for labeling antibody fragments with divalent or polyvalent radionuclides was developed. A bifunctional chelating agent, diethylenetriamine pentaacetic acid was covalently coupled, by an amide bond, to Fab fragments of antibodies to canine cardiac myosin. A stable chelate was then formed with indium-111, a nuclide that has appropriate half-life and energy characteristics for gamma imaging. Antibodies treated in this way retain their antigen-binding activity and are useful in locating myocardial infarcts in vivo.

Animals↗
PubMed1980

Renin-specific antibody for study of cardiovascular homeostasis.

Antiserum specific for purified canine renal renin was used to inhibit this enzyme in trained, conscious dogs. The antiserum did not affect blood pressure in sodium-replete dogs but decreased plasma renin activity and blood pressure in sodium-depleted animals. The antiserum also reduced blood pressure to control levels concomitant with suppression of plasma renin activity in uninephrectomized dogs with acute renovascular hypertension. These observations establish the role of the renin-angiotensin system in the maintenance of blood pressure in the sodium-depleted state as well as in the initiation of renovascular hypertension.

Animals↗
PubMed1980

A rapid and efficient procedure for the removal of nonviable cells from suspension culture of lymphoid cell lines.

The selective removal of nonviable cells from suspension cultures of lymphoid cell lines by buoyant density centrifugation at 400 x g on a Ficoll-Hypaque (FH) cushion is described. Viable cells collect at the medium-FH interface whereas nonviable cells pellet. This method has proven useful in separating viable from nonviable cells with human lymphoblastoid cell lines, rabbit lymphoid cell lines and mouse myeloma and hybridoma cell lines.

Animals↗
PubMed1980

Dissociation of renin--aldosterone and renal prostaglandin E during volume expansion induced by immersion in normal man.

The relationship of the renin--angiotensin--aldosterone axis with renal prostaglandin E is complex. Although studies have suggested that these two hormonal systems respond to experimental manipulations in a parallel manner, their interdependence has not been assessed fully during volume expansion. Since studies have demonstrated that in normal man the central hypervolaemia induced by water immersion to the neck produces a prompt and profound suppression of plasma renin activity and plasma aldosterone concentration without concomitant alteration of plasma composition, immersion afforded a unique opportunity to assess simultaneously the effects of central hypervolaemia on plasma renin activity, plasma aldosterone concentration and prostaglandin E excretion. 2. Seven normal subjects were studied twice while in balance on a diet containing 10 mmol of sodium/day, 100 mmol of potassium/day: with indomethacin administration (50 mg given every 6 h for five doses) and without indomethacin. Urinary prostaglandin E excretion was measured hourly and plasma renin activity and plasma aldosterone concentration at 30 min intervals. 3. Immersion was associated with a marked suppression of plasma renin activity (59 +/- 7%) and plasma aldosterone concentration (55 +/- 3%) with a return to pre-study values during the recovery hour. Concomitantly, urinary prostaglandin E excretion increased from 4.7 to a peak of 10.9 ng/min. Although administration of indomethacin lowered the basal rate of urinary prostaglandin E excretion and plasma renin activity, it did not prevent the subsequent augmentation of urinary prostaglandin E or the suppression of plasma renin activity and plasma aldosterone during the subsequent 4 h of immersion. 4. These results demonstrate a dissociation of renin--aldosterone and prostaglandin E during hypervolaemia and suggest that whereas prostaglandin E may constitute one of the major determinants of renin release clinically and experimentally, these two hormonal systems can be dissociated from each other in response to central volume expansion in man.

Adult↗
PubMed1980

Identification of renin and renin-like enzymes in rat brain by a renin-specific antibody.

1. The major angiotensin I-generating activity of rat brain extracts has a pH optimum different from that of renal renin and is not inhibited by renin specific antibody. 2. Affinity chromatography utilizing renin specific antibody, pepstatin and alpha-casein yielded fractions that resembled renal renin more closely with respect to antibody inhibition and pH optimum as well as an absence of the ability to hydrolyse haemoglobin. 3. We conclude that rat brain contains a host of enzymes with angiotensin I-generating activity including acid and neutral proteases and an enzyme with the immunochemical identity of renal renin. The biosynthetic origins of this renin-like protein remain uncertain.

Angiotensin I↗
PubMed1980

Specific inhibition of renin by an angiotensinogen analog: studies in sodium depletion and renin-dependent hypertension.

The angiotensin substrate analog Pro-His-Pro-Phe-His-Phe-Phe-Val-Tyr-Lys has no significant effect on blood pressure in sodium-replete monkeys (Macaca fascicularis) but blocks the pressor response to infused human renin. Pressor responses to angiotensin I and angiotensin II are not attenuated. In five studies in sodium-depleted monkeys, an infusion of 2 mg of the peptide per kg of body weight resulted in a reduction of mean arterial pressure (MAP) from 105 +/- 4 to 79 +/- 3 mm Hg, which is not significantly different from the response to 1 mg of the angiotensin I-converting enzyme inhibitor teprotide per kg. In uninephrectomized monkeys, inflation of a suprarenal aortic cuff caused an increase in MAP from 107 +/- 3 to 131 +/- 3 mm Hg. Infusion of 0.6 mg of the renin-inhibitory peptide per kg was followed by a return of blood pressure to 107 +/- 4 mm Hg--a depressor response similar to that observed with teprotide. This specific in vivo inhibitor of renin can now be applied to a wide variety of physiologic studies.

Angiotensin I↗
PubMed1980

Identification of an angiotensin receptor in rabbit renomedullary interstitial cells in tissue culture. Correlation with prostaglandin biosynthesis.

High affinity binding sites of angiotensin II (A II) have been characterized in rabbit renomedullary interstitial cells in tissue culture. Binding was rapid, specific, and saturable. Scatchard analysis of steady state saturation binding data at 22 degrees C indicated a single binding site with an equilibrium dissociation constant of 3.1 mM. The binding inhibition activities of analogues of angiotensin II correlated with their biological potencies: [Sar1, Ala8]A II greater than [des-Asp1]A II greater than greater than 3--8 hexapeptide. The biologically inactive 1--7 heptapeptide failed to displace the labeled angiotensin II from its binding sites. Angiotensin II directly stimulated prostaglandin E2 biosynthesis by the rabbit renomedullary interstitial cells in tissue culture. The concentration of angiotensin II that resulted in half maximal stimulation of prostaglandin E2 biosynthesis (6.5 nM) correlated with the concentration of angiotensin II that resulted in half maximal occupancy of binding sites (3.1 nM). The relative potency of [des-Asp1] A II as a stimulus for prostaglandin E2 biosynthesis was proportional to its relative binding inhibition activity. These findings demonstrate the biological significance of the angiotensin II binding sites in the rabbit renomedullary interstitial cells in tissue culture and suggest that the cells may provide a homogeneous model tissue for the study of the angiotensin receptor.

Angiotensin II↗
PubMed1980

Anti-alprenolol antibodies in the rabbit. A new probe for the study of beta-adrenergic receptor interaction.

We immunized rabbits with an antigen prepared by covalent linkage of alprenolol, a beta-adrenergic receptor antagonist, to bovine serum albumin. Competitive inhibition of [3H]dihydroalprenolol binding to antisera with a variety of unlabeled ligands revealed broad antibody specificity for beta-adrenergic antagonists and agonists. The antiserum was subjected to affintiy fractionation on hydroxybenzylpindolol-Sepharose 4B. Successive elution with 100 mM Tris HCl, 1M NaCl, 4 M LiBr, and 5 M guanidine yielded fractions with increasing affintiy for hydroxybenzylpindolol. The ligand-binding properties of these affinity-fractionated antibodies suggest that certain of these fractions recognize structural aspects of individual beta-adrenergic ligands which are irrelevant to their biological activity, whereas others can be used to distinguish shared functional properties, such as the ethanolamine side chain, within the structural heterogeneity of beta-adrenergic drugs. In particular, elution of hydroxybenzylpindolol-adsorbed antibody with (-)-propranolol allowed identification of an antibody fraction specific for the (-)-stereoisomer. Thus, affinity fractionation of antibodies raised against beta-adrenergic ligands can provide useful analogues for the further study of the recognition properties of the beta-adrenergic receptor.

Alprenolol↗