Archives of Biochemistry and Biophysics, Oct 1, 1988
Zinc binding to fibrinogen and fibrin was studied by two techniques. Scatchard analysis of ultraf... more Zinc binding to fibrinogen and fibrin was studied by two techniques. Scatchard analysis of ultrafiltration eluates reveals that fibrinogen has multiple Zn(II)-binding sites, KD (fibrinogen) = 18 microM; n = 6. The zinc content of the "collapsed" fibrin gel supernatant was also determined by atomic absorption spectroscopy and analyzed by a Scatchard plot (KD (fibrin) = 8 microM, n = 6). In other experiments, Zn(II) did not displace 45Ca(II) from fibrin. It appears that the binding of zinc to fibrinogen or fibrin is distinct from that of calcium, and that the zinc-binding characteristics of fibrinogen and fibrin are not significantly affected by the transformation of one into the other.
Exposure of albumin to Cu(II) (10-100 microM) and ascorbate (0.1-2 mM) results in extensive molec... more Exposure of albumin to Cu(II) (10-100 microM) and ascorbate (0.1-2 mM) results in extensive molecular modifications, indicated by decreased fluorescence and chain breaks. The rate of utilization of molecular oxygen and ascorbate as a function of Cu(II) concentration is non-linear at copper/albumin ratios of greater than 1. It appears that Cu(II) bound to the tightest albumin-binding site is less available to the ascorbate than the more loosely bound cation. SDS/polyacrylamide-gel electrophoresis reveals new protein bands corresponding to 50, 47, 22, 18 and 3 kDa. For such a cleavage pattern, relatively few (approximately 3) and rather specific chain breaks occurred. Repeated addition of portions of ascorbate to the albumin/Cu(II) mixture results in increased intensity of the new bands. The absence of Cu(II) or the presence of metal chelating agents is inhibitory. There was no evidence of intermolecular cross-linking or of the formation of insoluble, albumin-derived, material. A mech...
Fibrinogen is transformed into insoluble "neofibe" by reaction with up to 100 m... more Fibrinogen is transformed into insoluble "neofibe" by reaction with up to 100 microM Cu(II) and 1.5 mM ascorbate. The soluble peptides which are released during the reaction can be monitored by amino acid analysis and by measuring released keto-carbonyl (with DNPH). Immunologic characterization of the soluble peptides, with antibodies directed against fibrino-peptide A (FPA) clearly show the release of this epitope, optimally at 50 microM Cu(II). Anti-FPB gives no evidence of that epitope. However, N-terminal amino acid analyses reveals the presence of 3 peptides terminating in ALA (alpha chain FPA), GLU (beta chain FPB) and SER/ASP (unknown). The release of fibrinopeptides is interpreted within the context of a general mechanism for OH'-induced peptide chain cleavage via intermediate Schiff-base hydrolysis.
The biochemical basis of psychology and memory remain obscure. Psycho- and linguistic- analysis p... more The biochemical basis of psychology and memory remain obscure. Psycho- and linguistic- analysis provide few clues regarding the processes that occur in our heads. Here, I try to give an autobiographical account of the development of a scientific idea of how a mental process such as memory, can be encoded in the brain. My story began (~1980) with an account of how I started as a research biochemist working in blood coagulation, then became inspired by remarks in a book on linguistics (Noam Chomsky), to quote: • “Since the subject is a physical organism, the system attributed to it must have finite representations.” • “Symbol systems that fit different molds also have different neural representations…different physical representations.” • “An empirical argument must be brought to bear.” I formulated the concept of a “cation array” as the basis for the biochemical coding of information processing in the brain, as required for memory. The core idea was that the neuron employs various el...
Conflict of Interest: GM is a founder of MX Biotech Ltd., with the commercial goal to develop new... more Conflict of Interest: GM is a founder of MX Biotech Ltd., with the commercial goal to develop new classes of "memory materials" and devices. CG is emeritus professor of Chemistry at Hebrew University (Jerusalem), active in developing and patenting peptide-based tools for surgery and pharmacology. Notwithstanding, the ideas forwarded here by us both are scientifically genuine and presented in good faith, without commercial clouding of the concepts expressed herein.
Writing involves the transformation of ideas, emotions and sounds into graphic form. Ideagrams he... more Writing involves the transformation of ideas, emotions and sounds into graphic form. Ideagrams help clarify complex ideas and can be considered to be forms of stored memory. The emergence of chemistry from alchemy involved the adaptation of graphic notations to comprehend the invisible atomic and molecular structures of complex matter, and to spread and store that knowledge. This culminated in the molecular representations of chemical reactions and chemical compounds such as natural products, sugars, polymers, proteins and metal complexes. We review the development of textual and graphic notations of chemical structures from circa‐1700 to now. We present a Timeline that graphically summarizes the salient stages in the evolution of chemical notations (“chemography”) from alchemy, leading to modern “psycho‐chemistry”. The Timeline correlates the emergence of new chemical notations with crucial technological inventions.
International Journal of Dream Research, Apr 15, 2021
D I J o R Commentary "... each of us remembers and forgets in a pattern whose labyrinthing windin... more D I J o R Commentary "... each of us remembers and forgets in a pattern whose labyrinthing windings are an identification mark no less distinctive than a fingerprint". Philip Roth-American Pastoral "The pendulum of the mind oscillates between sense and nonsense, not between right and wrong." C.G. Jung, Memories, Dreams, Reflections "What takes us back to the past, are the memories. What brings us forward, is our dreams." Jeremy Irons, actor "Memories makes us who we are, and dreams makes us who we will become.
1. The neuron 2. Neural extracellular matrix (nECM) , the hydrogel around the cell which performs... more 1. The neuron 2. Neural extracellular matrix (nECM) , the hydrogel around the cell which performs as its "memory material". 3. Trace metals and neurotransmitters (NTs) distributed therein (dopants). The tripartite mechanism describes a chemical code for psychic states that is not linguistic, but presents the molecular correlates of the memory engram that render the neural synapse operative for the function of recall.
BACKGROUND "Every man's memory is his private literature."-Aldous Huxley. "I can only note that t... more BACKGROUND "Every man's memory is his private literature."-Aldous Huxley. "I can only note that the past is beautiful because one never realises an emotion at the time. It expands later, and thus we don't have complete emotions about the present, only about the past."-Virginia Woolf. Though pondered for millennia, neural mentation (cognition/ thought), memory and emotions experienced by all creatures, remain puzzling, psychic phenomena, examined by philosophers as well as physicists [1-7]. But "emotive memory" of any event is but a psychic shadow of a past experience. Notwithstanding, recalled emotions are what drive the organism's decision to react and act, as alluded to in some searching quotations from the American psychologist/philosopher William James (1884): "No shade of emotion is without bodily reverberations". " Emotion dissociated from all bodily feelings is unconceivable". "A disembodied human emotion is a non-entity". Emotions are expressed by simultaneous physiologic responses and psychic states. The literature on emotions, mentation and memory is enormous, contributed to by the likes of Aristotle,
This study was undertaken to determine if a plasma protease such as thrombin, a highly specific p... more This study was undertaken to determine if a plasma protease such as thrombin, a highly specific procoagulant enzyme that has numerous effects on platelets, endothelial cells, and smooth muscle cells, could mediate mast cell activation processes. Our results indicate that at near physiologic levels, thrombin can rapidly trigger mast cell degranulation without activating the 5-lipoxygenase system.
SummaryClinical observations have shown that heparin has antiinflammatory activities. The effect ... more SummaryClinical observations have shown that heparin has antiinflammatory activities. The effect of heparin on neutrophil chemotaxis was evaluated in vitro in the Boyden Chamber. This method enabled differentiation between the direct effects of heparin on neutrophil migration and locomotion, and its effects on chemotactic factors. Heparin inhibited both the random migration and directed locomotion of human neutrophils toward zymosan-activated serum (ZAS) and F-met-leu-phe (FMLP). Inhibition was found to be dependent on the concentrations of the heparin and of the chemotactic factors. No specific binding of heparin to the neutrophils could be demonstrated, and heparin’s inhibitory effects were eliminated by simple washing of the cells. When added directly to the chamber containing chemotactic factor, heparin inhibited the chemotactic activity of ZAS but not that of FMLP, suggesting a direct inhibitory effect against C5a, the principal chemotactic factor in ZAS.Experiments performed w...
SummaryThe mechanical properties of fibrin and protofibrin gels in the presence of physiologic le... more SummaryThe mechanical properties of fibrin and protofibrin gels in the presence of physiologic levels of Ca(II) and Zn(II) are described. As monitored with a thrombelastograph, Ca(II) (0.5–2 mM) increases the rate of development and the maximum level of gel elastic modulus (G) of fibrin and protofibrin gels. Zn(II) (10–50 μM) decreases the elastic modulus of those gels, even in the presence of a large excess of Ca(II). This contrasts with the ability of both divalent cations to increase fibrin and protofibrin gel turbidity. Unlike the turbidity or fibre thickness of fibrin and protofibrin gels, both of which are increased by these cations, gel elasticity is increased by Ca(II) but decreased by Zn(II). h is demonstrated that Ca(II) and Zn(II) modulate fibrin and protofibrin gels independently of one another, and that they have opposing effects on the mechanical properties of the gels. The disparity between the visual (turbidity, TEM) and the mechanical (elasticity) properties of (pro...
Background: The standard view of blood coagulation is based on a mechanism whereby cascade intera... more Background: The standard view of blood coagulation is based on a mechanism whereby cascade interactions of clotting factors generate thrombin, which converts soluble fibrinogen into an insoluble clot. Objective: Review the modalities by which soluble fibrinogen transforms into an insoluble matrix, the basis of blood coagulation. New concept: An alternate process is operative that can transform fibrinogen, based on reactions with free radicals. Such could be generated by the release of ascorbate by activated platelets. Ions of multivalent metals, such as Cu+2 or Fe+2 bound to fibrinogen, react with the ascorbate (a reductant in a Fenton reaction) to generate H2O2 and reactive oxygen species. Alternately, γ-irradiation which generates H2O2 could generate such species. Supportive evidence and references are cited. Conclusion: An expanded blood clotting schema is presented that incorporates the classic (via thrombin) as well as alternate (free radical) pathways by which fibrinogen can be converted into an insoluble clot. This new schema is discussed within the context of γ-irradiation or dietary ascorbate as instigants of free-radical induced clotting events, of particular relevance to airplane pilots, divers, submariners, astronauts and patients not responding to classic anticoagulation (heparin, Coumadin) therapy.
Fibrinogen has the potential of being used as a material to harvest and grow normal mesenchymal c... more Fibrinogen has the potential of being used as a material to harvest and grow normal mesenchymal cells (fibroblasts, endothelial cells) or to trap cancer cells from a suspension with blood as a potential circulatory trap.Insoluble fibrinogen particles (iFP) were prepared from commercial Cohn fraction I paste (source: Kedrion). The sized iFP (~60-180 µm) were not soluble in physiologic buffers, exhibited a density of 1.2 ± 0.02, and did not aggregate or clump when mixed with whole blood or thrombin, but were degraded in lytic solutions.Cell culture studies indicated that the iFP could be used to harvest, expand and transfer normal, mammalian, attachment-dependent cells, notably fibroblasts and stem cells from bone marrow, as well as numerous cancer lines. Cells attached to iFP underwent logarithmic growth kinetics and could be transferred without trypsinization. Transplanted cancer cells-on-iFP generated characteristic tumors and retained their surface marker (by Western immuno-blot). An iFP 'cell-affinity' batch column was shown to trap MCF-7 cancer cells in the presence of red blood cells (RBCs) or serum.The scalable process for fabricating iFP retained the cell attachment properties of native fibrinogen. The results indicate that iFP has the potential to be used as a 3D cell culture matrix, and possibly to trap cancer cells from blood.
Medical & Biological Engineering & Computing, 2006
The clotting time (CT) of fibrinogen mixed with thrombin decreased, then increased with increasin... more The clotting time (CT) of fibrinogen mixed with thrombin decreased, then increased with increasing fibrinogen levels. By contrast, log CT decreased monotonically with respect to the log level of activating enzyme (thrombin or reptilase). Here, the CT was determined over a large range of fibrinogen concentration (to 100 mg ml(-1)) at a fixed level of enzyme. A new parameter, [Fib]min, the minimal fibrinogen concentration required for thrombin or reptilase-instigated phase change (coagulation), was determined as [Fib]min = 0.2 +/- 0.05 microM fibrinogen. A dynamic simulation program (Stella) was employed to organize simulations based on simple and complex coagulation mechanisms, which generated CT values. The successful simulation aimed at forming [Fib]min and "recognized" the binding of unreacted fibrinogen with intermediate fibrin protofibrils. The "virtual data" mimicked the biphasic experimental CT values over a wide range of concentrations. Fibrinogen appeared to act in three modalities: as a thrombin substrate; as a precursor of fibrin; and as a competitor for fibrin protofibrils. The optimized simulation may provide a basis for predicting CT in more complex systems, such as pathological plasmas or whole blood or at high concentrations encountered with fibrin sealant.
Archives of Biochemistry and Biophysics, Oct 1, 1988
Zinc binding to fibrinogen and fibrin was studied by two techniques. Scatchard analysis of ultraf... more Zinc binding to fibrinogen and fibrin was studied by two techniques. Scatchard analysis of ultrafiltration eluates reveals that fibrinogen has multiple Zn(II)-binding sites, KD (fibrinogen) = 18 microM; n = 6. The zinc content of the "collapsed" fibrin gel supernatant was also determined by atomic absorption spectroscopy and analyzed by a Scatchard plot (KD (fibrin) = 8 microM, n = 6). In other experiments, Zn(II) did not displace 45Ca(II) from fibrin. It appears that the binding of zinc to fibrinogen or fibrin is distinct from that of calcium, and that the zinc-binding characteristics of fibrinogen and fibrin are not significantly affected by the transformation of one into the other.
Exposure of albumin to Cu(II) (10-100 microM) and ascorbate (0.1-2 mM) results in extensive molec... more Exposure of albumin to Cu(II) (10-100 microM) and ascorbate (0.1-2 mM) results in extensive molecular modifications, indicated by decreased fluorescence and chain breaks. The rate of utilization of molecular oxygen and ascorbate as a function of Cu(II) concentration is non-linear at copper/albumin ratios of greater than 1. It appears that Cu(II) bound to the tightest albumin-binding site is less available to the ascorbate than the more loosely bound cation. SDS/polyacrylamide-gel electrophoresis reveals new protein bands corresponding to 50, 47, 22, 18 and 3 kDa. For such a cleavage pattern, relatively few (approximately 3) and rather specific chain breaks occurred. Repeated addition of portions of ascorbate to the albumin/Cu(II) mixture results in increased intensity of the new bands. The absence of Cu(II) or the presence of metal chelating agents is inhibitory. There was no evidence of intermolecular cross-linking or of the formation of insoluble, albumin-derived, material. A mech...
Fibrinogen is transformed into insoluble "neofibe" by reaction with up to 100 m... more Fibrinogen is transformed into insoluble "neofibe" by reaction with up to 100 microM Cu(II) and 1.5 mM ascorbate. The soluble peptides which are released during the reaction can be monitored by amino acid analysis and by measuring released keto-carbonyl (with DNPH). Immunologic characterization of the soluble peptides, with antibodies directed against fibrino-peptide A (FPA) clearly show the release of this epitope, optimally at 50 microM Cu(II). Anti-FPB gives no evidence of that epitope. However, N-terminal amino acid analyses reveals the presence of 3 peptides terminating in ALA (alpha chain FPA), GLU (beta chain FPB) and SER/ASP (unknown). The release of fibrinopeptides is interpreted within the context of a general mechanism for OH'-induced peptide chain cleavage via intermediate Schiff-base hydrolysis.
The biochemical basis of psychology and memory remain obscure. Psycho- and linguistic- analysis p... more The biochemical basis of psychology and memory remain obscure. Psycho- and linguistic- analysis provide few clues regarding the processes that occur in our heads. Here, I try to give an autobiographical account of the development of a scientific idea of how a mental process such as memory, can be encoded in the brain. My story began (~1980) with an account of how I started as a research biochemist working in blood coagulation, then became inspired by remarks in a book on linguistics (Noam Chomsky), to quote: • “Since the subject is a physical organism, the system attributed to it must have finite representations.” • “Symbol systems that fit different molds also have different neural representations…different physical representations.” • “An empirical argument must be brought to bear.” I formulated the concept of a “cation array” as the basis for the biochemical coding of information processing in the brain, as required for memory. The core idea was that the neuron employs various el...
Conflict of Interest: GM is a founder of MX Biotech Ltd., with the commercial goal to develop new... more Conflict of Interest: GM is a founder of MX Biotech Ltd., with the commercial goal to develop new classes of "memory materials" and devices. CG is emeritus professor of Chemistry at Hebrew University (Jerusalem), active in developing and patenting peptide-based tools for surgery and pharmacology. Notwithstanding, the ideas forwarded here by us both are scientifically genuine and presented in good faith, without commercial clouding of the concepts expressed herein.
Writing involves the transformation of ideas, emotions and sounds into graphic form. Ideagrams he... more Writing involves the transformation of ideas, emotions and sounds into graphic form. Ideagrams help clarify complex ideas and can be considered to be forms of stored memory. The emergence of chemistry from alchemy involved the adaptation of graphic notations to comprehend the invisible atomic and molecular structures of complex matter, and to spread and store that knowledge. This culminated in the molecular representations of chemical reactions and chemical compounds such as natural products, sugars, polymers, proteins and metal complexes. We review the development of textual and graphic notations of chemical structures from circa‐1700 to now. We present a Timeline that graphically summarizes the salient stages in the evolution of chemical notations (“chemography”) from alchemy, leading to modern “psycho‐chemistry”. The Timeline correlates the emergence of new chemical notations with crucial technological inventions.
International Journal of Dream Research, Apr 15, 2021
D I J o R Commentary "... each of us remembers and forgets in a pattern whose labyrinthing windin... more D I J o R Commentary "... each of us remembers and forgets in a pattern whose labyrinthing windings are an identification mark no less distinctive than a fingerprint". Philip Roth-American Pastoral "The pendulum of the mind oscillates between sense and nonsense, not between right and wrong." C.G. Jung, Memories, Dreams, Reflections "What takes us back to the past, are the memories. What brings us forward, is our dreams." Jeremy Irons, actor "Memories makes us who we are, and dreams makes us who we will become.
1. The neuron 2. Neural extracellular matrix (nECM) , the hydrogel around the cell which performs... more 1. The neuron 2. Neural extracellular matrix (nECM) , the hydrogel around the cell which performs as its "memory material". 3. Trace metals and neurotransmitters (NTs) distributed therein (dopants). The tripartite mechanism describes a chemical code for psychic states that is not linguistic, but presents the molecular correlates of the memory engram that render the neural synapse operative for the function of recall.
BACKGROUND "Every man's memory is his private literature."-Aldous Huxley. "I can only note that t... more BACKGROUND "Every man's memory is his private literature."-Aldous Huxley. "I can only note that the past is beautiful because one never realises an emotion at the time. It expands later, and thus we don't have complete emotions about the present, only about the past."-Virginia Woolf. Though pondered for millennia, neural mentation (cognition/ thought), memory and emotions experienced by all creatures, remain puzzling, psychic phenomena, examined by philosophers as well as physicists [1-7]. But "emotive memory" of any event is but a psychic shadow of a past experience. Notwithstanding, recalled emotions are what drive the organism's decision to react and act, as alluded to in some searching quotations from the American psychologist/philosopher William James (1884): "No shade of emotion is without bodily reverberations". " Emotion dissociated from all bodily feelings is unconceivable". "A disembodied human emotion is a non-entity". Emotions are expressed by simultaneous physiologic responses and psychic states. The literature on emotions, mentation and memory is enormous, contributed to by the likes of Aristotle,
This study was undertaken to determine if a plasma protease such as thrombin, a highly specific p... more This study was undertaken to determine if a plasma protease such as thrombin, a highly specific procoagulant enzyme that has numerous effects on platelets, endothelial cells, and smooth muscle cells, could mediate mast cell activation processes. Our results indicate that at near physiologic levels, thrombin can rapidly trigger mast cell degranulation without activating the 5-lipoxygenase system.
SummaryClinical observations have shown that heparin has antiinflammatory activities. The effect ... more SummaryClinical observations have shown that heparin has antiinflammatory activities. The effect of heparin on neutrophil chemotaxis was evaluated in vitro in the Boyden Chamber. This method enabled differentiation between the direct effects of heparin on neutrophil migration and locomotion, and its effects on chemotactic factors. Heparin inhibited both the random migration and directed locomotion of human neutrophils toward zymosan-activated serum (ZAS) and F-met-leu-phe (FMLP). Inhibition was found to be dependent on the concentrations of the heparin and of the chemotactic factors. No specific binding of heparin to the neutrophils could be demonstrated, and heparin’s inhibitory effects were eliminated by simple washing of the cells. When added directly to the chamber containing chemotactic factor, heparin inhibited the chemotactic activity of ZAS but not that of FMLP, suggesting a direct inhibitory effect against C5a, the principal chemotactic factor in ZAS.Experiments performed w...
SummaryThe mechanical properties of fibrin and protofibrin gels in the presence of physiologic le... more SummaryThe mechanical properties of fibrin and protofibrin gels in the presence of physiologic levels of Ca(II) and Zn(II) are described. As monitored with a thrombelastograph, Ca(II) (0.5–2 mM) increases the rate of development and the maximum level of gel elastic modulus (G) of fibrin and protofibrin gels. Zn(II) (10–50 μM) decreases the elastic modulus of those gels, even in the presence of a large excess of Ca(II). This contrasts with the ability of both divalent cations to increase fibrin and protofibrin gel turbidity. Unlike the turbidity or fibre thickness of fibrin and protofibrin gels, both of which are increased by these cations, gel elasticity is increased by Ca(II) but decreased by Zn(II). h is demonstrated that Ca(II) and Zn(II) modulate fibrin and protofibrin gels independently of one another, and that they have opposing effects on the mechanical properties of the gels. The disparity between the visual (turbidity, TEM) and the mechanical (elasticity) properties of (pro...
Background: The standard view of blood coagulation is based on a mechanism whereby cascade intera... more Background: The standard view of blood coagulation is based on a mechanism whereby cascade interactions of clotting factors generate thrombin, which converts soluble fibrinogen into an insoluble clot. Objective: Review the modalities by which soluble fibrinogen transforms into an insoluble matrix, the basis of blood coagulation. New concept: An alternate process is operative that can transform fibrinogen, based on reactions with free radicals. Such could be generated by the release of ascorbate by activated platelets. Ions of multivalent metals, such as Cu+2 or Fe+2 bound to fibrinogen, react with the ascorbate (a reductant in a Fenton reaction) to generate H2O2 and reactive oxygen species. Alternately, γ-irradiation which generates H2O2 could generate such species. Supportive evidence and references are cited. Conclusion: An expanded blood clotting schema is presented that incorporates the classic (via thrombin) as well as alternate (free radical) pathways by which fibrinogen can be converted into an insoluble clot. This new schema is discussed within the context of γ-irradiation or dietary ascorbate as instigants of free-radical induced clotting events, of particular relevance to airplane pilots, divers, submariners, astronauts and patients not responding to classic anticoagulation (heparin, Coumadin) therapy.
Fibrinogen has the potential of being used as a material to harvest and grow normal mesenchymal c... more Fibrinogen has the potential of being used as a material to harvest and grow normal mesenchymal cells (fibroblasts, endothelial cells) or to trap cancer cells from a suspension with blood as a potential circulatory trap.Insoluble fibrinogen particles (iFP) were prepared from commercial Cohn fraction I paste (source: Kedrion). The sized iFP (~60-180 µm) were not soluble in physiologic buffers, exhibited a density of 1.2 ± 0.02, and did not aggregate or clump when mixed with whole blood or thrombin, but were degraded in lytic solutions.Cell culture studies indicated that the iFP could be used to harvest, expand and transfer normal, mammalian, attachment-dependent cells, notably fibroblasts and stem cells from bone marrow, as well as numerous cancer lines. Cells attached to iFP underwent logarithmic growth kinetics and could be transferred without trypsinization. Transplanted cancer cells-on-iFP generated characteristic tumors and retained their surface marker (by Western immuno-blot). An iFP 'cell-affinity' batch column was shown to trap MCF-7 cancer cells in the presence of red blood cells (RBCs) or serum.The scalable process for fabricating iFP retained the cell attachment properties of native fibrinogen. The results indicate that iFP has the potential to be used as a 3D cell culture matrix, and possibly to trap cancer cells from blood.
Medical & Biological Engineering & Computing, 2006
The clotting time (CT) of fibrinogen mixed with thrombin decreased, then increased with increasin... more The clotting time (CT) of fibrinogen mixed with thrombin decreased, then increased with increasing fibrinogen levels. By contrast, log CT decreased monotonically with respect to the log level of activating enzyme (thrombin or reptilase). Here, the CT was determined over a large range of fibrinogen concentration (to 100 mg ml(-1)) at a fixed level of enzyme. A new parameter, [Fib]min, the minimal fibrinogen concentration required for thrombin or reptilase-instigated phase change (coagulation), was determined as [Fib]min = 0.2 +/- 0.05 microM fibrinogen. A dynamic simulation program (Stella) was employed to organize simulations based on simple and complex coagulation mechanisms, which generated CT values. The successful simulation aimed at forming [Fib]min and "recognized" the binding of unreacted fibrinogen with intermediate fibrin protofibrils. The "virtual data" mimicked the biphasic experimental CT values over a wide range of concentrations. Fibrinogen appeared to act in three modalities: as a thrombin substrate; as a precursor of fibrin; and as a competitor for fibrin protofibrils. The optimized simulation may provide a basis for predicting CT in more complex systems, such as pathological plasmas or whole blood or at high concentrations encountered with fibrin sealant.
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