Terminal-modified polylysine-based chelating polymers: highly efficient coupling to antibody with minimal loss in immunoreactivity

Slinkin, M. A.; Klibanov, Alexander L.; Торчилин, В. П.

doi:10.1021/bc00011a009

Cited by 41 publications

(16 citation statements)

References 24 publications

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“…With 1.1 moles of Cy5.5 per mole of protein, labeling occurs at multiple lysine residues but was detectable at either of two lysine residues distal from the phospholipid‐binding plane of the protein. Owing to the sensitivity of annexin V to modification, more highly modified forms might best be accomplished by using a single‐point polymer attachment strategy19 or genetically modified annexin V 20. In addition, inactive, “overlabled” forms of dye–annexin Vs (Cy5.5 and Cy7) provide ideal controls for unspecific probe binding or accumulation, since many of their physical properties are similar to the dye–annexin Vs which bind apoptotic cells.…”

Section: Resultsmentioning

confidence: 99%

Optimal Modification of Annexin V with Fluorescent Dyes

2004

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The many uses of chemically modified annexin Vs necessitate an understanding of the optimal degree of modification and modification sites of the protein. When reacted with the N-hydroxysuccinimide ester of Cy5.5, annexin V with one modification per mole of protein retained its affinity for phosphatidylserine of apoptotic cells, whereas modification with two dyes per mole of protein caused a complete loss of activity. A tryptic digest LC/MS method was used to identify the modification sites as either of two closely spaced lysine residues, in position 286 or 290. The crystal structure indicated the location of these lysines was distal to the phosphatidylserine binding sites on annexin V. These results can be used to develop active or inactive fluorescent control annexin V proteins and to suggest strategies for attaining higher levels of modification with retention of bioactivity.

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Section: Resultsmentioning

confidence: 99%

Optimal Modification of Annexin V with Fluorescent Dyes

2004

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“…In this case, however, the lethal effect in mice decreased proportionally to PLA 2 activity. Some authors propose to assess different molar proportions of peptide: chelate or add some “linker” between the biomolecule and chelate to preserve the biological activity [ 20 , 21 ]. Use of “linkers” could be considered in future studies to understand the effect of bioconjugation process on enzymatic and lethal activity of β-NTx.…”

Section: Discussionmentioning

confidence: 99%

Biodistribution and Lymphatic Tracking of the Main Neurotoxin of Micrurus fulvius Venom by Molecular Imaging

Vergara

Castillo

Romero-Piña

et al. 2016

Toxins

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The venom of the Eastern coral snake Micrurus fulvius can cause respiratory paralysis in the bitten patient, which is attributable to β-neurotoxins (β-NTx). The aim of this work was to study the biodistribution and lymphatic tracking by molecular imaging of the main β-NTx of M. fulvius venom. β-NTx was bioconjugated with the chelator diethylenetriaminepenta-acetic acid (DTPA) and radiolabeled with the radionuclide Gallium-67. Radiolabeling efficiency was 60%–78%; radiochemical purity ≥92%; and stability at 48 h ≥ 85%. The median lethal dose (LD50) and PLA2 activity of bioconjugated β-NTx decreased 3 and 2.5 times, respectively, in comparison with native β-NTx. The immune recognition by polyclonal antibodies decreased 10 times. Biodistribution of β-NTx-DTPA-67Ga in rats showed increased uptake in popliteal, lumbar nodes and kidneys that was not observed with 67Ga-free. Accumulation in organs at 24 h was less than 1%, except for kidneys, where the average was 3.7%. The inoculation site works as a depot, since 10% of the initial dose of β-NTx-DTPA-67Ga remains there for up to 48 h. This work clearly demonstrates the lymphatic system participation in the biodistribution of β-NTx-DTPA-67Ga. Our approach could be applied to analyze the role of the lymphatic system in snakebite for a better understanding of envenoming.

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“…In addition, polymers with chelating agents have been used to label antibodies and proteins to reduce their loss of binding af nity (Slinkin, Klibanov, and Torchilin 1991;Torchilin et al 1987). Unmodi ed PEI has an inherent af nity to some metals (Spivakov, Geckeler, and Bayer 1985).…”

Section: Discussionmentioning

confidence: 99%

Biodistribution and Imaging of Polyethyleneimine-A Gene Delivery Agent

et al. 1999

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Polyethyleneimine (PEI) has been described as a potentially effective agent for gene delivery. To track the delivery of this gene vector, the biodistribution and imaging of PEI labeled with 111 Indium ( 111 In) was studied in Fischer 344 rats. PEI was conjugated with diethylenetriamine pentaacetic acid dianhydride (DTPA), dialyzed, and chelated with 111 In. Breast adenocarcinoma 13762 tumor cells were inoculated into the thighs of the rats. The rst group of rats (n = 3) were injected intravenously with 300 ¹g of the Indiumlabeled DTPA-polyethlyeimine ( 111 In-DTPA-PEI) (50 ¹Ci per rat) or 111 In-DTPA. These animals were imaged with a gamma camera with a medium energy parallel hole collimator at 5 min, 2 hr, and 24 hr postinjection. The percentage of uptake in tumor (region of interest) was quantitated by a computer image analyzer and expressed as a percentage of injected dose (%ID) per pixel. To further characterize the tissue distribution of 111 In-DTPA-PEI (300 l g, 10 ¹Ci per rat) and 111 In-DTPA (10 ¹Ci per rat), a second group of animals (n = 18) bearing breast tumors were studied with tissue uptake quanti ed at 2 hr, 24 hr, and 48 hr using a gamma counter. In addition, autoradiography was used to further characterize the distribution of the labeled polymer in two rats at 2 and 24 hr. From these studies, PEI was found to be rapidly cleared, primarily through the kidneys of the rats. In addition, the distribution of 111 In-DTPA-PEI was found to be signi cantly different from 111 In-DTPA with a higher tumor-to-blood ratio. These studies show that radio-labeled PEI may have potential as a gamma scintigraphy imaging agent and in tracking the delivery of genetic material.

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Terminal-modified polylysine-based chelating polymers: highly efficient coupling to antibody with minimal loss in immunoreactivity

Cited by 41 publications

References 24 publications

Optimal Modification of Annexin V with Fluorescent Dyes

Optimal Modification of Annexin V with Fluorescent Dyes

Biodistribution and Lymphatic Tracking of the Main Neurotoxin of Micrurus fulvius Venom by Molecular Imaging

Biodistribution and Imaging of Polyethyleneimine-A Gene Delivery Agent

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