scholarly journals Autoantibodies to phosphatidylethanolamine (PE) recognize a kininogen- PE complex

Blood ◽  
1995 ◽  
Vol 86 (8) ◽  
pp. 3083-3089 ◽  
Author(s):  
T Sugi ◽  
JA McIntyre
Keyword(s):  
Molecular Weight ◽  
Plasma Proteins ◽  
Enzyme Linked Immunosorbent Assay ◽  
Low Molecular Weight ◽  
High Molecular Weight Kininogen ◽  
Recurrent Spontaneous Abortions ◽  
Glycoprotein I ◽  
Column Eluate ◽  
Adult Bovine Serum ◽  
Elisa Plate

Demonstration of autoimmune antiphospholipid antibodies (aPA) to negatively charged phospholipids (PL) in an enzyme-linked immunosorbent assay (ELISA) requires the presence of certain phospholipid-binding plasma proteins, eg, beta 2-glycoprotein I. We found a requirement for plasma against the electrically neutral or zwitterionic phospholipid, phosphatidylethanolamine (PE). Two of these PE-binding plasma proteins were identified as high molecular weight kininogen (HMWK) and low molecular weight kininogen (LMWK). We studied anti-PE antibody (aPE) seropositive plasma from 13 patients with SLE and/or recurrent spontaneous abortions by using partially purified kininogens and kininogen binding proteins from adult bovine serum isolated by carboxymethyl (CM)-papain affinity chromatography. Eleven of 13 sera recognized a kininogen-PE complex and/or a kininogen-binding protein-kininogen-PE complex. Some aPE-positive patient sera were shown to recognize highly purified HMWK and LMWK by ELISA only when the kininogens were presented on a PE substrate. These aPE sera did not recognize PE, HMWK, or LMWK when they were presented independently as the sole antigens on the ELISA plates. Other aPE-positive sera that did not react with PE-bound HMWK or LMWK reacted with the CM-papain column eluate when it was bound to PE, which suggests that these aPE recognize factor XI or prekallikrein, which normally bind to HMWK. The aPE ELISA reactivity of two patient sera were inhibited by preincubation of the CM-papain column eluate in the ELISA plate. These data show that most aPE are not specific for PE but require the presence of certain PL-binding plasma proteins that are kininogens or proteins in complex with kininogens. Our studies indicate that aPE bind to different plasma proteins than those implicated in anionic PL, aPA ELISA reactivity.

scholarly journals Autoantibodies to phosphatidylethanolamine (PE) recognize a kininogen- PE complex

Blood ◽  
1995 ◽  
Vol 86 (8) ◽  
pp. 3083-3089 ◽  
Author(s):  
T Sugi ◽  
JA McIntyre
Keyword(s):  
Molecular Weight ◽  
Plasma Proteins ◽  
Enzyme Linked Immunosorbent Assay ◽  
Low Molecular Weight ◽  
High Molecular Weight Kininogen ◽  
Recurrent Spontaneous Abortions ◽  
Glycoprotein I ◽  
Column Eluate ◽  
Adult Bovine Serum ◽  
Elisa Plate

Abstract Demonstration of autoimmune antiphospholipid antibodies (aPA) to negatively charged phospholipids (PL) in an enzyme-linked immunosorbent assay (ELISA) requires the presence of certain phospholipid-binding plasma proteins, eg, beta 2-glycoprotein I. We found a requirement for plasma against the electrically neutral or zwitterionic phospholipid, phosphatidylethanolamine (PE). Two of these PE-binding plasma proteins were identified as high molecular weight kininogen (HMWK) and low molecular weight kininogen (LMWK). We studied anti-PE antibody (aPE) seropositive plasma from 13 patients with SLE and/or recurrent spontaneous abortions by using partially purified kininogens and kininogen binding proteins from adult bovine serum isolated by carboxymethyl (CM)-papain affinity chromatography. Eleven of 13 sera recognized a kininogen-PE complex and/or a kininogen-binding protein-kininogen-PE complex. Some aPE-positive patient sera were shown to recognize highly purified HMWK and LMWK by ELISA only when the kininogens were presented on a PE substrate. These aPE sera did not recognize PE, HMWK, or LMWK when they were presented independently as the sole antigens on the ELISA plates. Other aPE-positive sera that did not react with PE-bound HMWK or LMWK reacted with the CM-papain column eluate when it was bound to PE, which suggests that these aPE recognize factor XI or prekallikrein, which normally bind to HMWK. The aPE ELISA reactivity of two patient sera were inhibited by preincubation of the CM-papain column eluate in the ELISA plate. These data show that most aPE are not specific for PE but require the presence of certain PL-binding plasma proteins that are kininogens or proteins in complex with kininogens. Our studies indicate that aPE bind to different plasma proteins than those implicated in anionic PL, aPA ELISA reactivity.

Comparison of the Non-Specific Binding of Unfractionated Heparin and Low Molecular Weight Heparin (Enoxaparin) to Plasma Proteins

1993 ◽  
Vol 70 (04) ◽  
pp. 625-630 ◽  
Author(s):  
Edward Young ◽  
Benilde Cosmi ◽  
Jeffrey Weitz ◽  
Jack Hirsh
Keyword(s):  
Molecular Weight ◽  
Unfractionated Heparin ◽  
Plasma Proteins ◽  
Low Molecular Weight Heparin ◽  
Specific Binding ◽  
Anticoagulant Activity ◽  
Factor Xa ◽  
Low Molecular Weight ◽  
Heparin Binding ◽  
Fixed Amount

SummaryThe non-specific binding of anticoagulantly-active heparin to plasma proteins may influence its anticoagulant effect. We used low affinity heparin (LAH) essentially devoid of anti-factor Xa activity to investigate the extent and possible mechanism of this non-specific binding. The addition of excess LAH to platelet-poor plasma containing a fixed amount of unfractionated heparin doubled the anti-factor Xa activity presumably because it displaces anticoagulantly-active heparin from plasma proteins. Although dextran sulfates of varying molecular weights also increased the anti-factor Xa activity, less sulfated heparin-like polysaccharides had no effect. These findings suggest that the ability to displace active heparin from plasma protein binding sites is related to charge and may be independent of molecular size. In contrast to its effect in plasma containing unfractionated heparin, there was little augmentation in anti-factor Xa activity when LAH was added to plasma containing low molecular weight heparin (LMWH), indicating that LMWH binds less to plasma proteins than unfractionated heparin. This concept is supported by studies comparing the anticoagulant activity of unfractionated heparin and LMWH in plasma with that in buffer containing antithrombin III. The anti-factor Xa activity of unfractionated heparin was 2-fold less in plasma than in the purified system. In contrast, LMWH had identical anti-factor Xa activity in both plasma and buffer, respectively. These findings may be clinically relevant because the recovered anti-factor Xa activity of unfractionated heparin was 33% lower in plasma from patients with suspected venous thrombosis than in plasma from healthy volunteers. The reduced heparin recovery in patient plasma reflects increased heparin binding to plasma proteins because the addition of LAH augmented the anti-factor Xa activity. In contrast to unfractionated heparin, there was complete recovery of LMWH added to patient plasma and little increase of anti-factor Xa activity after the addition of LAH. These findings may explain why LMWH gives a more predictable dose response than unfractionated heparin.

Low Molecular Weight Heparin Therapy: Is Monitoring Needed?

1994 ◽  
Vol 72 (03) ◽  
pp. 330-334 ◽  
Author(s):  
B Boneu
Keyword(s):  
Molecular Weight ◽  
Clinical Trials ◽  
Plasma Proteins ◽  
Bleeding Risk ◽  
Heparin Therapy ◽  
Low Molecular Weight ◽  
Vein Thrombosis ◽  
Meta Analyses ◽  
Deep Vein ◽  
Initial Check

SummaryRecent meta-analyses indicate that low molecular weight heparins (LMWH) are more effective than unfractionated heparin (UH) in preventing and treating deep vein thrombosis. This article presents the arguments for and against the need for laboratory monitoring. At the present time, the only tests currently available for monitoring LMWH therapy are those which measure the anti Xa activity in the plasma. Due to lower binding to plasma proteins and to cell surfaces,the plasma anti Xa activity generated by a given dose of LMWH is more predictable than for UH.Some clinical trials suggest that LMWH delivered at the recommended dose expose the patient to less bleeding risk than UH. Several . meta-analyses indicate comparable risk while any overdose unaccept-ably increases the haemorrhagic risk. The lowest dose of LMWH still effective in treating established DVT is presently unknown; some reports indicate that inadequate doses of LMWH are associated with a lack of efficacy for prevention. An overview of the published clinical trials indicates that the LMWH dose has never been monitored for prevention of DVT. In the treatment of established DVT, several trials have been performed without any monitoring, while in others the dose was adapted to target a given anti Xa activity. These considerations suggest that in prevention of DVT, monitoring the dose is not required. In the treatment of established DVT, considering the haemorrhagic risk of LMWH, the risk of undertreating the patient and the absence of large clinical trials comparing the advantages of monitoring the dose or not, it might be useful to check anti Xa activity at least once at the beginning of the treatment but the need for this initial check remains to be established. Because a large proportion of patients will be in the desired range, dose adjustments will be far less frequent than for UH.

Neutralisation of Heparan Sulphate and Low Molecular Weight Heparin by Protamine

1985 ◽  
Vol 53 (01) ◽  
pp. 086-089 ◽  
Author(s):  
A R Hubbard ◽  
C A Jennings
Keyword(s):  
Molecular Weight ◽  
Plasma Proteins ◽  
Low Molecular Weight Heparin ◽  
Heparan Sulphate ◽  
Activated Partial Thromboplastin Time ◽  
Weight Basis ◽  
Low Molecular Weight ◽  
Protamine Sulphate ◽  
At Iii ◽  
Reference Preparation

SummaryThe neutralisation by protamine sulphate (PS) of heparan sulphate (HS), a low molecular weight heparin (LMWH), and a reference preparation of unfractionated heparin (UH), was studied by activated partial thromboplastin time (APTT) and anti-Xa clotting assays. UH was most easily neutralised in the APTT assay by PS (on a weight for weight basis), followed by LMWH and HS. The neutralisation of APTT activity by PS closely followed the loss of activity in the anti-Xa clotting assay, when plasma was used as the source of At III. When the anti-Xa clotting assay was carried out using purified At III in place of plasma, HS and LMWH were neutralised by much lower amounts of PS and resembled UH neutralisation more closely. Resistance of HS anti-Xa activity to PS neutralisation decreased with increasing plasma dilution. The presence of bovine albumin with purified At III concentrate increased the resistance of HS to PS neutralisation. It is concluded that PS binding to UH, HS and LMWH is probably related more to their degree of sulphation than molecular weight and that non-specific interactions between PS and plasma proteins inhibit the binding of PS to HS and LMWH.

Low molecular weight plasma proteins in the cerebrospinal fluid of children with hematological malignancies

1984 ◽  
Vol 12 (2) ◽  
pp. 131-136 ◽  
Author(s):  
M. B. Duggan ◽  
J. A. Whittaker ◽  
E. H. Cooper ◽  
C. C. Bailey ◽  
E. A. Robinson
Keyword(s):  
Molecular Weight ◽  
Cerebrospinal Fluid ◽  
Plasma Proteins ◽  
Hematological Malignancies ◽  
Low Molecular Weight

scholarly journals Cucumber Mosaic Virus, Tobacco Streak Virus, and Cucumber Mosaic Virus Satellite RNA Associated with Mosaic and Ringspot Symptoms in Ajuga reptans in Ohio

Plant Disease ◽  
1997 ◽  
Vol 81 (10) ◽  
pp. 1214-1214 ◽  
Author(s):  
J. R. Fisher ◽  
S. T. Nameth
Keyword(s):  
United States ◽  
Molecular Weight ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
The United States ◽  
Negative Control ◽  
Enzyme Linked Immunosorbent Assay ◽  
Low Molecular Weight ◽  
Streak Virus ◽  
Ajuga Reptans

Creeping bugleweed (Ajuga reptans L.) is a perennial ornamental commonly grown as a ground cover in temperate climates. Commercial samples of the A. reptans cultivars Royalty, var. Atropurpurea Bronze, Bronze Beauty, and Burgundy Glow showing mosaic and ringspot symptoms were tested for the presence of virus infection by direct antibody sandwich enzyme-linked immunosorbent assay (ELISA) and viral-associated double-stranded (ds) RNA analysis. Cucumber mosaic cucumovirus (CMV) was detected by ELISA and dsRNA analysis in symptomatic samples of all cultivars tested. ELISA values were considered positive if the absorbance values were twice the negative control. Negative control values were established with asymptomatic tissue of the cv. Bronze Beauty. Tobacco streak ilarvirus (TSV) was detected only by ELISA in symptomatic samples of all cultivars except Royalty. No dsRNA suggestive of TSV was detected. Alfalfa mosaic virus (AMV) was detected by ELISA and dsRNA analysis in symptomatic samples of all cultivars tested except Royalty and var. Atropurpurea Bronze. dsRNA analysis also indicated the presence of a low molecular weight, possible satellite (sat) RNA associated with all symptomatic and asymptomatic Royalty and var. Atropurpurea Bronze plants tested. Northern (RNA) blot analysis with a digoxigenin-labeled full-length clone of the (S) CARNA-5 (-) CMV satRNA (ATCC no. 45124) confirmed that the low molecular weight RNA associated with the Royalty and var. Atropurpurea Bronze cultivars was indeed CMV satRNA. Only AMV has been previously reported in A. reptans in the United States (1). This is the first report of CMV and its satRNA, as well as TSV, in A. reptans in the United States. Reference: (1) W. T. Schroeder and R. Provvidenti. Plant Dis. Rep. 56:285, 1972.

Investigations of the Immunoglobulin Subtype Transformation of Anti–Heparin-Platelet Factor 4 Antibodies During Treatment With a Low-Molecular-Weight Heparin (Clivarin) in Orthopedic Patients

2003 ◽  
Vol 127 (5) ◽  
pp. 584-588
Author(s):  
Sarfraz Ahmad ◽  
H. Peter Bacher ◽  
Michael R. Lassen ◽  
Debra A. Hoppensteadt ◽  
Helen Leitz ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Low Molecular Weight Heparin ◽  
Serotonin Release ◽  
Platelet Factor 4 ◽  
Enzyme Linked Immunosorbent Assay ◽  
Low Molecular Weight ◽  
Platelet Factor ◽  
Significant Difference ◽  
Antibody Titers ◽  
Orthopedic Patients

Abstract Context.—It is now widely accepted that the pathophysiology of heparin-induced thrombocytopenia (HIT) syndrome is mediated by the generation of a wide array of functional and molecularly heterogeneous anti–heparin-platelet factor 4 (AHPF4) antibodies that may mediate platelet and/or endothelial cell activation/destruction. Objective.—We investigated the differential prevalence and functionality of AHPF4 immunoglobulin subtypes (IgA, IgG, and IgM) in plasmas obtained from orthopedic patients immobilized with Plaster-Cast and treated with clivarin (a low-molecular-weight heparin) in comparison to a placebo for the prophylaxis of deep-vein thrombosis. Design and Methods.—Clivarin was administered subcutaneously at a fixed daily dosage of 1750 U without any adjustment or loading dosage. Citrated plasmas were obtained at baseline, at 10 to 14 days, and at postbrace procedure (5–12 weeks). An enzyme-linked immunosorbent assay (ELISA) was used to quantitate the AHPF4 antibody titers. The functionality of the ELISA-positive samples was determined by a 14C-serotonin release assay (SRA). Results.—In the ELISA test, 16 of 1073 samples (1.5%; 6 in clivarin and 10 in placebo groups) were positive for AHPF4 antibodies (mean optical density [OD] = 0.46 ± 0.02). None of the ELISA-positive samples for AHPF4 antibodies could mediate platelet activation responses as determined by the SRA (0%–3% serotonin release, P > .10, n = 16). Through differential immunoglobulin subtype analysis of the samples positive for (cumulative) AHPF4 antibodies, we determined that their relative prevalence in plasma were as follows: IgM (mean OD = 0.71 ± 0.13) > IgG (0.31 ± 0.08) > IgA (0.14 ± 0.02). Although there was no significant difference in the total antibody titers between clivarin and placebo groups, the antibody subtyping data showed conversion trends (ie, IgA [clivarin to placebo], IgG [placebo to clivarin], and IgM [clivarin to placebo]). Conclusion.—These observations indicate that even at reduced dosages, clivarin can shift the immunogenic up-regulation toward the IgG subpopulation; however, the IgG subtype is of a nonfunctional type of AHPF4 antibody and thus may not cause any HIT-related pathogenic responses.

Isolation and Characterization of Human Platelet β-Thromboglobulin

1975 ◽  
Author(s):  
D. S. Pepper ◽  
S. Moore ◽  
J. D. Cash
Keyword(s):  
Molecular Weight ◽  
Plasma Proteins ◽  
Human Platelet ◽  
Weight Fraction ◽  
Human Platelets ◽  
Low Molecular Weight ◽  
Purification Procedure ◽  
Isolation And Characterization

The thrombin released products from washed human platelets were separated by filtration on 4% agarose in 0.15 M NaCl. The high molecular weight PF4 complex was dissociated and re-chromatographed in 0.75 M NaCl. The low molecular weight fraction, including β thromboglobulin and a low MW anti-heparin was freed of plasminogen anti-activator by dissociation and chromatography in pH 3.5 pyridine acetic acid. The anti-activator was irreversibly denatured and albumin was removed in the void volume of the column. A more suitable purification procedure for recovery of all activities was affinity chromatography on heparin-agarose. The anti-activator was excluded and could be obtained free of plasma proteins by Sephadex G-200 chromatography. The βTG eluted at 0.3 M NaCl and the low MW anti-heparin at 1.5 M NaCl. The pure βTG (MW 36,000) was injected into rabbits and the resulting antiserum used to produce a radioimmunoassay for the release reaction in vivo.

Comparison of exopolysaccharides from mucoid and nonmucoid strains of Clavibacter michiganensis subspecies sepedonicus

1993 ◽  
Vol 39 (3) ◽  
pp. 291-296 ◽  
Author(s):  
Paul J. Henningson ◽  
Neil C. Gudmestad
Keyword(s):  
Molecular Weight ◽  
High Performance ◽  
Size Exclusion ◽  
Enzyme Linked Immunosorbent Assay ◽  
Low Molecular Weight ◽  
Neutral Sugar ◽  
Clavibacter Michiganensis ◽  
Serological Reaction ◽  
Molecular Weights ◽  
Exclusion Chromatography

The exopolysaccharides produced by six strains of Clavibacter michiganensis ssp. sepedonicus were isolated and purified by liquid chromatography. Neutral sugar composition and molecular weights were determined for each polysaccharide fraction, using gas chromatography and high-performance size-exclusion chromatography. The serological reaction of each fraction was tested using enzyme-linked immunosorbent assay. Exopolysaccharide from nonmucoid strains contained only low molecular weight polysaccharides (1.5 × 103 to 1.1 × 104). Exopolysaccharide from mucoid and intermediate strains could be separated into low (4.0 × 103 to 1.1 × 104) molecular weight and high (5.0 × 105 to 1.6 × 106) molecular weight fractions. High molecular weight polysaccharides were composed almost exclusively of galactose, glucose, and fucose. The ratios of these sugars were highly variable among strains. Low molecular weight polysaccharides were primarily composed of galactose with significant and varying amounts of glucose, rhamnose, mannose, and ribose. All polysaccharide fractions except one, produced by a nonmucoid strain, reacted in the immunoassay test.Key words: exopolysaccharide, polysaccharide, Clavibacter, michiganensis, sepedonicus.

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