Biomedical subjects

V R Muzykantov

Publications and source records attributed to V R Muzykantov.

At least 37 records · Page 2Linked to original sources

PubMed1996

Target-sensitive immunoerythrocytes: interaction of biotinylated red blood cells with immobilized avidin induces their lysis by complement.

Red blood cells (RBC) coated with antibody (immunoerythrocytes) may be useful for drug targeting. Previously we have developed a methodology for avidin (streptavidin)-mediated attachment of biotinylated antibodies (b-Ab) to biotinylated RBC (B-RBC). We have observed that binding of avidin to B-RBC in suspension leads to their complement-mediated lysis by autologous serum. In the present work we have studied the interaction of B-RBC, which are not complement susceptible, with immobilized avidin and their consequent susceptibility to lysis by complement. B-RBC adhered tightly to avidin-coated surfaces and were rendered susceptible to lysis by autologous serum. A long biotin ester provided more effective binding of the B-RBC to immobilized avidin and greater lysis by complement, than a short biotin ester. Based on these results, we have hypothesized that targeting of serum-stable drug-loaded B-RBC attained by step-wise administration of b-Ab and streptavidin may provide target-sensitive lysis of B-RBC. To confirm this hypothesis, we have studied b-Ab and streptavidin mediated targeting of B-RBC to immobilized antigen. Step-wise addition of biotinylated antibody, avidin or streptavidin and b-RBC caused specific binding of B-RBC to immobilized antigen and their subsequent lysis by autologous serum. Therefore, our results obtained in an in vitro model demonstrate that B-RBC might be used for targeting and local release of drug.

Aminocaproates↗

PubMed1996

Portal hypertension enhances endotoxin-induced intercellular adhesion molecule 1 up-regulation in the rat.

BACKGROUND & AIMS: Liver disease or portosystemic shunting enhances th e sensitivity to endotoxin. The aim of this study was to investigate whether intercellular adhesion molecule 1 (ICAM-1) expression in response to endotoxin may be dysregulated in an animal model of portal hypertension. METHODS: Portal hypertension was induced by partial portal vein ligation. Sham-operated animals served as controls. ICAM-1 expression was measured using radiolabeled antibodies under baseline conditions or 5 hours after treatment with either endotoxin or recombinant tumor necrosis factor (TNF). Immunoreactive plasma TNF was also measured. RESULTS: Under baseline conditions, ICAM-1 expression in all organs studied was similar in portal-hypertensive and sham-operated rats. ICAM-1 up-regulation after a high dose of endotoxin (5 mg/kg) was similar in both groups of animals. However, portal-hypertensive animals showed a significantly higher ICAM-1 expression in response to low doses of endotoxin (0.1-10 microgram/kg). The response to a low (but not a high) dose of recombinant TNF was also significantly enhanced in portal-hypertensive animals. In addition, portal-hypertensive rats had higher plasma TNF levels after treatment with endotoxin or recombinant TNF. CONCLUSIONS: Portal hypertension induces an exaggerated ICAM-1 up-regulation in response to endotoxin, which is related to an increased production and decreased clearance of the cytokine.

Analysis of Variance↗

PubMed1996

Endothelial cells internalize monoclonal antibody to angiotensin-converting enzyme.

We investigated the fate of MAb 9B9, a monoclonal antibody to angiotensin-converting enzyme (ACE), which binds to endothelium both in vitro and in vivo. Using cultured human umbilical vein endothelial cells (HUVEC) and isolated perfused rat lungs (IPL), we demonstrated specific and saturable binding of 125I-labeled MAb 9B9 at 4 degrees C [affinity constant (Kd) = 20-50 nM, maximal number of binding sites (Bmax) = 1.5-3.0 x 10(5) sites/cell]. When 125I-MAb 9B9 was bound to HUVEC at 37 degrees C, only 40% of cell-associated radioactivity was acid elutable, suggesting antibody internalization. This was confirmed by finding that 1) the amount of MAb 9B9 uptake at 37 degrees C was higher than at 4 degrees C both in HUVEC and IPL; 2) binding of 125I-labeled streptavidin with HUVEC and IPL pretreated with biotinylated MAb 9B9 (b-MAb 9B9) was diminished in a temperature- and time-dependent fashion at 37 degrees C; and 3) b-MAb 9B9 bound to HUVEC at 37 degrees C was found intracellularly by ultrastructural analysis using streptavidin gold. Intracellular 125I-MAb 9B9 was found in microsomal fractions of lung homogenate from IPL and after intravenous (iv) injections in rats. Degradation of internalized MAb 9B9 was minimal, since > 90% of cell-associated 125I label remained precipitable by trichloracetic acid in HUVEC, IPL, and in vivo. Autoradiography of sodium dodecyl sulfate-polyacrylamide gel electrophoresis of lung homogenates made as late as several days after iv injections of 125I-MAb 9B9 in rats demonstrated a predominant band above 140 kDa. These data indicate that endothelial cells either in vitro or in vivo internalize the ACE ligand MAb 9B9 without significant intracellular degradation. Therefore MAb 9B9 may be useful for selective intracellular delivery of drugs to the pulmonary vascular endothelium after systemic administration.

Animals↗

PubMed1996

Attachment of antibody to biotinylated red blood cells: immuno-red blood cells display high affinity to immobilized antigen and normal biodistribution in rats.

Streptavidin-mediated attachment of biotinylated antibodies (b-Ab) to biotinylated red blood cells (b-RBC) is useful for preparation of immuno-red blood cells, a prospective vehicle for drug targeting. However, streptavidin (SA) induces lysis of extensively biotinylated RBC by complement due to cross-linking and inactivation of RBC complement regulators. To reduce cross-linking of RBC membrane proteins, we utilized mild biotinylation of RBC with 20 microM biotin ester (b20-RBC). SA effectively binds to rat b20-RBC (10(5) SA molecules/cell) and provides for following attachment of 5 x 10(4) molecules of b-IgG/SA per b20-RBC. By in vitro assay, b-Ab/SA/b20-RBC were stable in fresh rat serum. Serum-stable immuno-red blood cells (b-Ab/SA/b20-RBC) specifically bound to antigen-coated surfaces, but not to BSA-coated surfaces. Biodistribution of 51Cr-labelled b-Ab/SA/b20-RBC in rats was similar to that of control RBC, with no indication of lysis in vivo. These results suggest b-Ab/SA/b20-RBC may be explored as a vehicle for drug targeting.

Animals↗

PubMed1996

Targeting of antibody-conjugated plasminogen activators to the pulmonary vasculature.

Thrombolytic therapy has not been widely used for pulmonary embolism due to less than optimal results with conventional plasminogen activators. We propose a new approach to deliver plasminogen activators to the luminal surface of the pulmonary vasculature to potentially improve dissolution of pulmonary thromboemboli. Our previous studies have documented that a monoclonal antibody (mAb) to angiotensin-converting enzyme (anti-angiotensin-converting enzyme mAb 9B9) accumulates in the lungs of various animal species after systemic administration. We coupled 125I-labeled biotinylated plasminogen activators (single-chain urokinase plasminogen activator, tissue-type plasminogen activator and streptokinase) to biotinylated mAb 9B9, using streptavidin as a cross-linker. The fibrinolytic activity of plasminogen activators was not changed significantly by either biotinylation or by coupling to streptavidin. Antibody-conjugated plasminogen activators bind to the antigen immobilized in plastic wells and provide lysis of fibrin clots formed in these wells. Therefore, antibody-conjugated plasminogen activators bound to their target antigen retain their capacity to activate plasminogen. One hour after i.v. injection of mAb 9B9-conjugated radiolabeled biotinylated single-chain urokinase plasminogen activator, biotinylated tissue-type plasminogen activator or biotinylated-streptokinase in rats, the level of radiolabel was 7.4 +/- 0.8, 5.9 +/- 0.4 and 3.6 +/- 0.4% of injected dose/g (ID/g) of lung tissue vs. 0.5 +/- 0.01, 0.3 +/- 0.01 and 0.6 +/- 0.3% ID/g after injection of the same activators conjugated with control mouse IgG (P < .01 in all cases). Injection of mAb 9B9-conjugated radiolabeled plasminogen activator led to its rapid pulmonary uptake with a peak value 6.2 +/- 1.2% ID/g attained 3 hr after injection. One day later, 2.2 +/- 0.5% of the injected radioactivity was found per gram of lung tissue, although the blood level was 0.13 +/- 0.03% ID/g (lung/blood ratio 16.7 +/- 0.3). Therefore, conjugation of plasminogen activators with anti-angiotensin-converting enzyme mAb 9B9 provides their specific targeting to and prolonged association with the pulmonary vasculature. These results provide a basis for study of the local pulmonary fibrinolysis by mAb 9B9-conjugated plasminogen activators.

Animals↗

PubMed1995

Enhanced complement susceptibility of avidin-biotin-treated human erythrocytes is a consequence of neutralization of the complement regulators CD59 and decay accelerating factor.

Biotinylation of erythrocytes (E) followed by avidin cross-linking at specific sites has been suggested as a novel means of drug delivery. Upon avidin cross-linking, biotinylated E become complement-activating and highly susceptible to complement lysis, thus bringing about release of entrapped drug. We set out to examine the mechanisms of this biotin-avidin-induced lytic susceptibility, focusing on the effects of biotinylation and avidin cross-linking on the major E complement regulatory molecules, decay accelerating factor (DAF) and CD59. We demonstrate here that biotinylation of E, which does not render them complement activating, partially inhibits DAF but has little effect on CD59. Subsequent cross-linking with avidin causes complete inhibition of DAF and near complete loss of CD59 activity. Following cross-linking, DAF and CD59 become associated in high molecular mass avidin-containing complexes on the membrane. Incorporation of physiological amounts of CD59 into the membranes of biotinylated and avidin cross-linked E is sufficient to render these cells resistant to complement lysis whereas incorporation of DAF has relatively little effect. An understanding of the molecular mechanisms underlying complement susceptibility of biotin-avidin treated E should allow a rational design of strategies for drug delivery using E or other large, potentially complement-activating carriers.

Antibodies↗

PubMed1995

The functional effects of biotinylation of anti-angiotensin-converting enzyme monoclonal antibody in terms of targeting in vivo.

The effect of modification with biotin N-hydroxysuccinimide ester of mouse monoclonal antibody to angiotensin-converting enzyme, anti-ACE Mab 9B9, on its targeting to endothelial cells has been studied in vitro and in vivo. By in vitro assay, Mab 9B9 biotinylated at a biotin/IgG molar ratio in reaction mixture (B/IgG ratio) of 0.7-2.2 bound streptavidin monovalently and retained antigen-binding capacity. Mab 9B9 biotinylated at a B/IgG ratio of 20 and higher bound streptavidin polyvalently. Extensive biotinylation (B/IgG ratio of 60 and higher) led to dramatic reduction of Mab 9B9 Ag-binding capacity and to reduction of Mab 9B9 recognition by goat polyclonal antibody to mouse IgG. Radiolabeled Mab 9B9 biotinylated at a B/IgG ratio of 6 (b6-Mab 9B9) bound effectively to cultured vascular endothelium, with affinity characteristics similar to nonbiotinylated Mab 9B9. Endothelial cells internalized both Mab 9B9 and b6-Mab 9B9 to the same extent (60% internalization at 3 h incubation at 37 degrees C). Degradation of cell surface-associated Mab 9B9 or b6-Mab 9B9 was very low (< 1% as measured by TCA solubility of radiolabel). In contrast, degradation of internalized b6-Mab 9B9 was more profound than that of Mab 9B9 (20 +/- 3% vs 6 +/- 1%, P < 0.01). After injection in rats, radiolabeled b6-Mab 9B9 had a biodistribution pattern similar to that of radiolabeled Mab 9B9. Both preparations effectively accumulated in the lung (15-20% of injected dose/g of tissue vs 2% of injected dose/g of blood).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

Attachment of biotinylated antibody to red blood cells: antigen-binding capacity of immunoerythrocytes and their susceptibility to lysis by complement.

A biotinylated monoclonal antibody (mAb) to human IgM (b-anti-IgM) has been attached to human red blood cells (RBC) by two different approaches. The first method is performed with biotinylated RBC (b-RBC) and involves stepwise binding of streptavidin (SA) to b-RBC followed by addition and binding of specific b-anti-IgM or b-IgG. b-RBC were prepared with differing input levels of biotin N-hydroxysuccinimide ester (BNHS). At moderate BNHS levels (100 microM) the resulting b-RBC (designated b4-RBC) bound 50,000 molecules of b-IgG after treatment with SA. However, at high BNHS levels (> 1000 microM) the resulting b-RBC bound b-IgG poorly, presumably due to multivalent binding of each SA to several biotins in close proximity on the RBC. b-RBC prepared at high BNHS inputs (but not b4-RBC) were lysed by serum plus SA. Stepwise attachment of b-anti-IgM to SA-coated b4-RBC allows binding of up to 6 x 10(4) molecules of b-anti-IgM/RBC. The second method is based on attachment of b-anti-IgM to RBC via CR1, the primate RBC complement receptor. The SA-biotin system is used to prepare bi-specific mAb complexes (heteropolymers) in which a biotinylated mAb to CR1 is cross-linked with b-anti-IgM via SA. Binding of these heteropolymers to RBC via CR1 is specific and saturable and can facilitate binding of up to 2500 molecules of b-anti-IgM/RBC.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

Interaction of mAb to angiotensin-converting enzyme (ACE) with antigen in vitro and in vivo: antibody targeting to the lung induces ACE antigenic modulation.

We previously described that mAb to angiotensin-converting enzyme (ACE), mAb 9B9, accumulates in the rat lungs after systemic injection. In the present work we have documented that mAb 9B9 cross-reacts with human, monkey, rat, cat and hamster ACE, while other ACE antibodies did not cross-react with the rat, cat and hamster enzyme. Anti-ACE mAb 3A5 and I2H5 inhibit human ACE in vitro, while mAb 9B9 does not inhibit ACE activity. Radiolabeled mAb 9B9, but not other antibodies, accumulates selectively in rat, cat and hamster lungs after systemic administration. No accumulation of mAb 9B9 has been observed in hamster kidney, while hamster kidney ACE activity is higher than that in the lung. mAb 9B9 does not induce complement-mediated injury to cultured endothelial cells. No pathological changes were detected in organs of animals after mAb 9B9 injection (10-100 mg/kg). However, injection of these amounts of mAb 9B9 leads to a decrease in ACE activity in the lung homogenates and an increase in serum. In cultured human endothelial cells treatment with mAb 9B9 increases ACE activity in cell medium and decreases in cell lysates. Therefore, while mAb 9B9 does not kill endothelial cells, at high dose it may induce ACE shedding from the cell. The results obtained support the potential of anti-ACE mAb 9B9 for targeting to the lung and for investigations of the pulmonary endothelium.

Animals↗

PubMed1994

Immunotargeting of streptavidin to the pulmonary endothelium.

UNLABELLED: We have observed previously that monoclonal antibody to angiotensin-converting enzyme (Mab 9B9) accumulates selectively in the lung after intravenous injection. The objective of the present work is the development of a universal system for targeting of drug or radiolabel to the lung, using biotinylated Mab 9B9 and streptavidin. METHODS: Mab 9B9 was biotinylated with biotin succinimide ester (b-Mab 9B9), while streptavidin (SA) was radiolabeled with 125I. Interaction between b-Mab 9B9 and SA has been estimated in solid-phase radioassay. Radiolabeled SA was conjugated with b-Mab 9B9 or with b-IgG and injected intravenously in rats or perfused in isolated rat lungs. RESULTS: Radiolabeled b-Mab 9B9 biotinylated at biotin-to-antibody molar ratio 10 (b-Mab 9B9) retains its ability to accumulate in rat lungs after intravenous injection. Radiolabeled SA conjugated with b-Mab 9B9 accumulates in the lung tissue in perfused isolated rat lungs. About 20% of injected SA accumulates in the rat lung 1 hr after intravenous injection (localization ratio is 20, immunospecificity of the conjugate pulmonary uptake is 70). As compared with conjugate injection, stepwise intravenous injection of b-Mab 9B9 and radiolabeled SA leads to a marked reduction of SA pulmonary uptake. Maximal pulmonary uptake of Mab 9B9 has been observed 2-3 hr after intravenous injection, while 24 hr later, radioactivity in the lung was markedly reduced. In contrast to radiolabeled Mab 9B9 alone, radiolabeled SA conjugated with b-Mab 9B9 was retained in the lung for at least 48 hr. In concert with effective blood clearance of the conjugate, its prolonged lung retention leads to a marked increase in its lung-to-blood ratio: 80 for SA-b-Mab 9B9 versus 15-20 for Mab 9B9. CONCLUSION: Conjugation of Mab 9B9 with streptavidin enhances selective pulmonary uptake of the preparation, providing a background for intrapulmonary immunotargeting of various biotinylated agents.

Animals↗

PubMed1993

Interaction of avidin-carrying red blood cells with nucleated cells.

In vivo application of red blood cells (RBC) modified with avidin-biotin complex has been suggested recently for various purposes. However, avidin attachment to RBC alters their biocompatibility. Thus, it has been described that avidin-carrying biotinylated RBC were lysed by the complement. In the present work interaction between avidin-carrying RBC and nucleated cells has been examined. It was found that attachment of avidin, but not streptavidin, to RBC led to binding of avidin-carrying RBC to nucleated cells. Adhesiveness of nucleated cells for avidin-carrying RBC varied for different types of nucleated cells. The strongest adhesion was observed with human fibroblasts and rat Kupffer cells, while rat liver endothelial cells were practically non-adhesive for avidin-carrying RBC of corresponding species. In contrast with avidin (streptavidin)-induced lysis by the complement, avidin-induced adhesion was independent of temperature, the presence of divalent ions and mode of avidin attachment. Polyanions (dextran sulphate and heparin) efficiently inhibited the adhesion presumably due to interaction with the membrane-bound avidin. Polyanions to a much lesser extent inhibited lysis of avidin-carrying RBC, which might be a result of their interaction with the complement components. Polycations also blocked adhesion of avidin-carrying RBC to nucleated cells, presumably due to interaction with negatively charged cell-surface components. Therefore, attachment of avidin to RBC alters their biocompatibility, due to both high positive charge of avidin and the cross-linking of biotinylated membrane proteins.

Animals↗

PubMed1993

Avidin attachment to biotinylated human neutrophils induces generation of superoxide anion.

The influence of biotinylation and subsequent attachment of avidin on generation of superoxide anion by human neutrophils was studied. Biotinylation of human neutrophils with succinimide ester of biotin does not reduce superoxide generation in response to activation with phorbol myristate acetate (PMA) and formyl peptide (FMLP). Addition of avidin to biotinylated, but not native, leukocytes induces generation of superoxide anion. The kinetics and level of superoxide generation by biotinylated neutrophils in response to addition of avidin were quite similar to those in response to activation with FMLP. The avidin sugar moiety and charge were not involved in superoxide generation, since streptavidin was also active. Both avidin- and PMA-induced superoxide generation were independent of the extracellular calcium, while FMLP-induced superoxide generation was dependent on the presence of calcium in solution. Therefore, interaction of avidin with biotinylated components of the neutrophil membrane alters functional activity of this cell and might induce 'activation-like' reaction of leukocytes.

1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium ↗

PubMed1993

Avidin attachment to biotinylated amino groups of the erythrocyte membrane eliminates homologous restriction of both classical and alternative pathways of the complement.

Lysis of avidin-coated biotinylated sheep red blood cells (RBC) via the classical pathway of homologous (sheep) and heterologous (guinea pig) complement has been studied. The minimal surface density of avidin inducing antibody-dependent lysis via the classical pathway is smaller than that inducing antibody-independent lysis via the alternative pathway. Heterologous lysis via the classical pathway does not depend on the mode of avidin attachment: both biotinylation of membrane amino groups and insertion of biotinyl-lipid into the membrane provide the same lysis of avidin-coated RBCs by guinea pig serum in the presence of anti-avidin antibody. Avidin-free sheep RBC sensitized with hemolytic anti-RBC antibody were lysed by guinea pig, but not by sheep serum, confirming high efficiency of homologous restriction of the complement. However, avidin-coated RBCs were lysed by homologous serum in the presence of anti-avidin antibody at low surface density of avidin attached. The elimination of the homologous restriction depends on the mode of avidin attachment: biotinylation of membrane amino groups provides antibody-mediated lysis via the classical pathway of homologous complement, while insertion of biotinyl-lipid does not provide lysis.

Amines↗

PubMed1993

Avidin attachment to red blood cells via a phospholipid derivative of biotin provides complement-resistant immunoerythrocytes.

Preincubation of red blood cells (RBC) in an aqueous dispersion of biotin-phosphatidylethanolamine (biotin-PE) provides binding sites for avidin on the surface of these cells (up to 5 x 10(5) avidin molecules per cell). Previously we have shown that biotin covalently attached to the surface of RBC by a chemical reaction with biotin N-hydroxysuccinimide ester permits attachment of avidin to these cells, resulting in the activation of the alternative pathway of complement with subsequent cell lysis. However, avidin attached to RBC via biotin-PE did not cause complement activation. This is not due to the stabilizing action of biotin-PE. In contrast, various phospholipids, including biotin-PE, enhance the lysis of RBC induced by hemolytic antibodies via the classical complement pathway. The potential of avidin-coated RBC to act as activators of the complement alternative pathway depends on the method of biotin attachment to RBC. Complement-resistant avidin-coated RBC can specifically bind biotinylated antibodies. These immunoerythrocytes effectively and specifically bind to the antigen-coated surface and are not lysed by complement even in the presence of soluble antigen. These data extend the possible applications of immunoerythrocytes in drug targeting.

Animals↗

PubMed1993

Tannin-mediated attachment of avidin provides complement-resistant immunoerythrocytes that can be lysed in the presence of activator of complement.

It was shown previously that avidin attachment to biotinylated erythrocytes induces their lysis by homologous complement via the alternative pathway. This phenomenon hinders the use of avidin-coated immunoerythrocytes as carriers for drug targeting. In the present work we demonstrated that attachment of avidin to erythrocytes via the cross-linking agent tannin does not induce their lysis by complement. Tannization allows attachment of about 5 x 10(5) molecules of avidin per erythrocyte, which is comparable to the value obtained after treatment with biotin esters. In contrast to biotinylated avidin-coated erythrocytes, tannized avidin-coated erythrocytes were not lysed by complement. Tannization itself does not reduce the erythrocyte sensitivity to lysis by complement in the presence of activators of the complement (hemolytic antibody or activators of the alternative pathway). Therefore, the avidin-induced lysis by complement depends on the mode of avidin attachment to erythrocyte. Complement-resistant tannized erythrocytes coated with avidin bind biotinylated immunoglobulins (to 7 x 10(4) molecules per cell), suggesting that tannization might be used for the preparation of complement-resistant immunoerythrocytes.

Animals↗

PubMed1993

Sublethal doses of exogenous hydrogen peroxide prime human neutrophils to formyl peptide.

Hydrogen peroxide (H2O2) is an oxidative agent important in inflammation and ischemia. Neutrophils (PMNs) are a main source of H2O2 in the inflammatory focus. However, after recruitment into the inflammatory or ischemic zone of the heart, the PMN itself might serve as a target for exogenous H2O2. In the present work we found that PMNs are very resistant to the cytotoxic action of H2O2 (LD50 for PMNs is about 30-50 mM, whereas for endothelial cells it is about 200-300 microM). Unexpectedly, treatment of PMNs by H2O2 at a sublethal dose of 10 mM leads to a subsequent increase in the generation of superoxide anion in response to the chemoattractant peptide FMLP (twofold increase in O2- generation 30 min after treatment by H2O2 as compared with nontreated control cells). H2O2 itself does not induce O2- generation by PMNs. Therefore, any H2O2 that accumulated in the inflammatory or ischemic zone might alter the functional activity of PMNs and prime them to subsequent agonist activation.

Centrifugation, Density Gradient↗

PubMed1993

[Tannin-mediated attachment of avidin to erythrocytes does not cause their lysis by complement].

It was shown previously that avidin attachment to biotinylated erythrocytes induced their lysis by a homologous complement via an alternative pathway. This phenomenon hindered the use of avidin-coated immuno-erythrocytes as carriers for drug targeting. In the present work it has been demonstrated that avidin attachment to erythrocytes via a cross-linking reagent (tannin) does not induce any lysis by the complement. Tannization provides an attachment of up to 5 x 10(5) avidin molecules per erythrocyte which is commensurate with the value obtained after treatment with biotin esters. However, in contrast with biotinylated avidin-coated erythrocytes tannized cells are not lysed by the complement, while tannization itself does not diminish the erythrocyte sensitivity to lysis by the complement in the presence of activators (hemolytic antibody or activators of the alternative pathway). The avidin-induced lysis by the complement depends on the mode of avidin attachment to erythrocytes. Complement-resistant avidin-coated tannized erythrocytes bind biotinylated immunoglobulins and may therefore be used as carriers for drug targeting. The use of hemolytic antibody in biotinylated immunoglobulins attached to avidin-coated erythrocytes provides their controlled lysis by a complement activated via a classical pathway.

Avidin↗

PubMed1992

Avidin-induced lysis of biotinylated erythrocytes by homologous complement via the alternative pathway depends on avidin's ability of multipoint binding with biotinylated membrane.

It was reported that avidin and streptavidin induce lysis of prebiotinylated red blood cells via the alternative pathway of both homologous and heterologous complement. Both of these proteins have four biotin-binding sites, providing a polyvalent interaction with biotinylated components of the erythrocyte membrane. We have compared the effects of mono- and multipoint avidin attachment on the sensitivity of biotinylated erythrocytes to lysis by the complement system. In the presence of anti-avidin antibody, avidin-bearing biotinylated erythrocytes were rapidly lysed by heterologous serum. This lysis was independent from the mode of avidin attachment, implying that complement activation by the classical pathway triggered by interaction between C1 and avidin-bound antibody on the erythrocyte surface is independent from the avidin's ability of polyvalent (multipoint) binding with biotinylated membrane components. In the absence of anti-avidin antibody, biotinylated erythrocytes bearing polyvalently attached avidin were lysed by homologous complement better than cells bearing avidin, which possesses reduced ability for multipoint binding with biotinylated erythrocyte. Two independent approaches to reduce avidin's ability of multipoint binding were used: decrease in surface density of biotin on the erythrocyte membrane and blockage of biotin-binding sites of avidin. Both methods result in reduced lysis of avidin-bearing erythrocytes as compared with erythrocytes bearing an equal amount of polyvalent-bound avidin. Thus the activation of homologous complement via the alternative pathway depends on avidin's ability to 'cross-link' to the biotinylated components of the erythrocyte membrane.

Animals↗