Name: an optimized protocol for the efficient radiolabeling of gold nanoparticles by using a 125i-labeled azide prosthetic group
Uploaded: 2016-10-10T15:00:00.000+00:00
Duration: 7 min 23 s
Description: Watch this Scientific Journal Video about An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group at JoVE.com

This work was supported by grants from the National Research Foundation of Korea, funded by the government of the Republic of Korea, (Grant nos. 2012M2B2B1055245 and 2012M2A2A6011335) and by the RI-Biomics Center of Korea Atomic Energy Research Institute.

זהירות: צורת החמצון של יוד רדיואקטיבי היא תנודתי למדי ויש לטפל עם מגינים להוביל נאותים בקבוקוני עופרת. כל צעדי radiochemical צריכות להתבצע במנדף פחם-מסונן מאוורר היטב, ואת הפרוצדורות צריכות להיות פיקוח על ידי זיהוי התקני רדיואקטיביות. 1. הכנת הכימיקלים והמחסנית הפוך שלב לסינתזה של 125 I-שכותרתו יזיד <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> הכנת ריאגנטים בתמיסה <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> ממסי 1 מ&quot;ג של המבשר יזיד (2) ב 150 μl אתנול אבסולוטי (איור 1). הערה: הליך סינטטי מפורט המבשר יזיד (2) דווחה בעיתון הקודם 22. </li><li style=";text-align:right;direction:rtl"> ממיסים T chloramine 1 מ&quot;ג ב 20 μl של חיץ מלוחים פוספט 1x (pH = 7.4). </li><li style=";text-align:right;direction:rtl"> ממיסים metabisulfite נתרן 2 מ&quot;ג ב 20 μl H 2 O. </li></ol></li><li style=";text-align:right;direction:rtl"> preparation של המחסנית <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לשטוף את המחסנית tC18 עם אתנול אבסולוטי 10 מ&quot;ל ואחריו 10 מ&quot;ל H 2 O. אין לייבש את המטריצה ​​של מחסנית עם אוויר. </li></ol></li></ol> 2. Radiosynthesis של 125 הקבוצה התותבת תזיד שכותרתו לי <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> תגובת Radioiodination של המבשר <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> מוסיפים את פתרון מבשר אזיד (1 מ&quot;ג ב 150 μl של אתנול אבסולוטי) וחומצה אצטית (10 μl) לצינור 1.5 מ&quot;ל microcentrifuge. </li><li style=";text-align:right;direction:rtl"> להוסיף 150 MBq של [125 I] נאה ב 0.1 M NaOH (50 μl) לתערובת התגובה. </li><li style=";text-align:right;direction:rtl"> הוספת פתרון T chloramine (1 מ&quot;ג ב 20 μl של חיץ מלוחים פוספט 1x) ולסגור את תערובת התגובה המכיל microcentrifuge צינור. </li><li style=";text-align:right;direction:rtl"> דגירה תערובת התגובה בטמפרטורת החדר למשך 15 דקות עד השלמת התגובה radioiodination. </li><li style=";text-align:right;direction:rtl"> הוספת פתרון metabisulfite נתרן (2 מ&quot;ג ב 20 μl H 2 O) לתערובת התגובהכדי להרוות את התגובה radioiodination. </li><li style=";text-align:right;direction:rtl"> לסגת 0.2 μl של המוצר הגולמי ואז לדלל אותו עם 100 μl של פתרון (H 2 O / CH 3 CN, 1: 1) עבור ניתוח HPLC. הערה: עבור כל הניסויים HPLC, השתמש 0.1% חומצה פורמית המכיל H 2 O (ממס) ו -0.1% אצטוניטריל המכילים חומצה פורמית (B ממס) כמו eluents. </li><li style=";text-align:right;direction:rtl"> נתח את המוצר הגולמי מדולל באמצעות הפוכה פאזיים אנליטיים רדיו-HPLC (C18 הפוכה שלב בטור; קצב הזרימה: 1 מ&quot;ל / דקה; שיפוע eluent: 20% B ממס עבור 0-2 דקות, 20-80% ב ממס עבור 2-22 דקות, B ממס 80-100% עבור 22-23 דקות, ולאחר 100% B ממס עבור 23-28 דקות; זמן השמירה: 16.4 דק &#39;) (איור 2). </li></ol></li><li style=";text-align:right;direction:rtl"> טיהור של המוצר הגולמי עם HPLC preparative הערה: ספק מספיק שריון עופרת סביב חלקי HPLC כגון מזרק, עמודה, גלאי, בקבוקונים אוספים, ואת המכל שבו הקולחים נאספו. <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לסגת הדואר תערובת תגובה כולו לתוך בקבוקון HPLC. יש לשטוף את צינור התגובה עם אצטוניטריל (0.5 מ&quot;ל) ולהוסיף השטיפה לתוך אותו בקבוקון ההזרקה. לדלל את הפתרון שנאספו עם H 2 O (1 מ&quot;ל). </li><li style=";text-align:right;direction:rtl"> להזריק את המוצר הגולמי על רדיו-HPLC preparative (C18 הפוכה שלב בטור; קצב הזרימה: 10 מ&quot;ל / דקה; שיפוע eluent: 20% B ממס עבור 0-2 דקות, 20-80% ב ממס עבור 2-22 דקות, 80-100% B ממס עבור 22-23 דקות, ולאחר 100% B ממס עבור 23-28 דקות). </li><li style=";text-align:right;direction:rtl"> אספו את שיא רדיואקטיבי המייצג 125 יזיד שכותרתו אני (1) (R t בתנאי HPLC אלה הוא 17.8-18.8 דקות) במבחנת זכוכית (איור 2). </li><li style=";text-align:right;direction:rtl"> מדוד את תשואת radiochemical של השבר באמצעות כיל רדיואקטיביות מינון על פי הפרוטוקול של היצרן. </li><li style=";text-align:right;direction:rtl"> להזריק את המוצר המטוהר תתפס-HPLC רדיו אנליטיים באמצעות באותם התנאים HPLC לקביעת טוהר radiochemical של המוצר. </li></ol></li><li style=";text-align:right;direction:rtl"> מיצוי שלב מוצק של המוצר <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לדלל את שבריר המכיל את המוצר הרצוי (1) עם 40 מ&quot;ל טהור H 2 O. </li><li style=";text-align:right;direction:rtl"> מוסיף את הפתרון המדולל לתוך מחסנית tC18 preconditioned. </li><li style=";text-align:right;direction:rtl"> לשטוף את המחסנית עם H מ&quot;ל 15 נוספים 2 O. </li><li style=";text-align:right;direction:rtl"> Elute המוצר (1) לכוד בתוך מחסנית עם 2 מ&quot;ל אצטון לתוך בקבוקון זכוכית 10 מ&quot;ל כי הוא מוגן על ידי מגן עופרת. מדוד את הרדיואקטיביות של מוצר eluted באמצעות כיל רדיואקטיביות מינון על פי הפרוטוקול של היצרן. הערה: sulfoxide דימתיל (DMSO) או אתנול אבסולוטי יכול לשמש גם עבור elution של המוצר מהמחסנית. כ 5-10% של רדיואקטיביות בדרך כלל מקלות על המחסנית, המוצר radiolabeled הנותרים לא ניתן eluted מלא באמצעות כמויות גדולות של ממיס אורגני. </li><li style=";text-align:right;direction:rtl"> לאדות את אצטון עם זרם של חנקן או גז ארגון. </li><li style=";text-align:right;direction:rtl"> ממיסים את מחדשsidue עם DMSO (100-200 μl) עבור השלב radiolabeling הבא. </li></ol></li></ol> סינתזת 3. DBCO-group-מצומדות חלקיקי זהב <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> שינוי פני שטח של זהב 13 ננומטר בגודל חלקיקים עם פוליאתילן גליקול קבוצה-DBCO המכיל <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> כן חלקיקי זהב נתרן מיוצב-ציטראט (3) (= גודל הממוצע 13 ננומטר) על פי דו&quot;ח קודם 24. </li><li style=";text-align:right;direction:rtl"> להוסיף בתמיסה מימית של 20 Tween (1 מ&quot;מ, 1.5 מ&quot;ל) אל חלקיקי זהב מיוצב ציטרט (10 ננומטר, 15 מ&quot;ל). לנער את הפתרון עבור 20 דקות על שייקר מסלולית. </li><li style=";text-align:right;direction:rtl"> להוסיף בתמיסה מימית של תיאול פוליאתילן גליקול DBCO המכילים-קבוצה (משקל מולקולרי ממוצע = 5000, 100 מיקרומטר, 1.5 מ&quot;ל). לנער את פתרון 2 שעות על שייקר מסלולית. </li></ol></li><li style=";text-align:right;direction:rtl"> טיהור של חלקיקי זהב DBCO-שונה-group <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לטהר את חלקיקי זהב DBCO-שונה-קבוצה (4) &lt;/ Strong&gt; על ידי צנטריפוגה רצוף (11,400 XG, 15 דק &#39;x 3). </li><li style=";text-align:right;direction:rtl"> למזוג supernatant ולהוסיף מים טהורים עבור resuspension של כדורי ננו-חלקיקים מזהב. </li></ol></li></ol> 4. Radiolabeling של ננו-חלקיקי זהב DBCO-שונה-קבוצה דרך התגובה לחץ חופשי הנחושת <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> סינתזה של 125 ננו-חלקיקי זהב שכותרתו לי באמצעות 125 I-שכותרתו יזיד (1) <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> כן פתרון מרוכז של חלקיקי זהב DBCO-שונה-קבוצה באמצעות צנטריפוגה (11,400 XG, 15 דק &#39;), ולהתאים את הריכוז של חלקיקי זהב עד 2 מיקרומטר. </li><li style=";text-align:right;direction:rtl"> להוסיף 4.1 MBq של אזיד 125 שכותרתו אני (1) ב DMSO (5 μl) כדי השעיה של חלקיקי זהב (4) (2 מיקרומטר, 50 μl). </li><li style=";text-align:right;direction:rtl"> דגירת תערובת התגובה וכתוצאה מכך ב -40 מעלות צלזיוס במשך 60 דקות. </li><li style=";text-align:right;direction:rtl"> משיכת דוגמא מתמיסה בסיסית (0.2 μl) מהמוצר הגולמי ולהחיל אותו על שיתוף סיליקהכרומטוגרפיה בשכבה דקה ated (TLC) צלחת. </li><li style=";text-align:right;direction:rtl"> לפתח את צלחת TLC באמצעות אתיל יצטט כשלב נייד. </li><li style=";text-align:right;direction:rtl"> מניחים את צלחת TLC על סורק רדיו-TLC ולהפעיל את הסורק כדי לפקח על התגובה radiolabeling (איור 3) על פי פרוטוקול של היצרן. </li></ol></li><li style=";text-align:right;direction:rtl"> טיהור של המוצר הגולמי <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לטהר את תערובת התגובה המכיל את חלקיקי זהב 125 שכותרתו לי (4) על ידי צנטריפוגה (11,400 XG, 15 דק &#39;). </li><li style=";text-align:right;direction:rtl"> למזוג supernatant ולהוסיף מים טהורים עבור resuspension של כדורי ננו-חלקיקים מזהב. </li><li style=";text-align:right;direction:rtl"> משיכת דוגמא מתמיסה בסיסית (0.2 μl) מהמוצר המטוהר ולהחיל אותו על צלחת TLC מצופה סיליקה. </li><li style=";text-align:right;direction:rtl"> לפתח את צלחת TLC באמצעות אתיל יצטט כשלב הנייד. </li><li style=";text-align:right;direction:rtl"> מניחים את צלחת TLC על סורק רדיו-TLC ולהפעיל את הסורק כדי לקבוע את התשואה radiochemical וטוהר radiochemical של 125 gol שכותרתו ליחלקיקי ד (4) (איור 3) על פי הפרוטוקול של היצרן. </li></ol></li></ol>

<ol>
	<li>Jewett, J. C., Bertozzi, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cu-free+Click+Cycloaddition+Reactions+in+Chemical+Biology.">Cu-free Click Cycloaddition Reactions in Chemical Biology.</a> Chem. Soc. Rev. 39, 1272-1279 (2010).</li><li>Debets, M. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioconjugation+with+Strained+Alkenes+and+Alkyne.">Bioconjugation with Strained Alkenes and Alkyne.</a> Acc. Chem. Res. 44, 805-815 (2011).</li><li>Sletten, E. M., Bertozzi, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=From+Mechanism+to+Mouse:+A+Tale+of+Two+Bioorthogonal+Reactions.">From Mechanism to Mouse: A Tale of Two Bioorthogonal Reactions.</a> Acc. Chem. Res. 44, 666-676 (2011).</li><li>Koo, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioorthogonal+Cu-Free+Click+Chemistry+in+vivo+for+Tumor-Targeted+Delivery+of+Nanoparticles.">Bioorthogonal Cu-Free Click Chemistry in vivo for Tumor-Targeted Delivery of Nanoparticles.</a> Angew. Chem. Int. Ed. 51, 11836-11840 (2012).</li><li>Chang, P. V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Copper-Free+Click+Chemistry+in+Living+Animals.">Copper-Free Click Chemistry in Living Animals.</a> Proc. Natl. Acad. Sci. U. S. A. 107, 1821-1826 (2010).</li><li>Bostic, H. E., Smith, M. D., Poloukhtine, A. A., Popik, V. V., Best, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Membrane+Labeling+and+Immobilization+via+copper-free+Click+Chemistry.">Membrane Labeling and Immobilization via copper-free Click Chemistry.</a> Chem. Commun. 48, 1431-1433 (2012).</li><li>Someya, T., Ando, A., Kimoto, M., Hirao, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Site-Specific+Labeling+of+RNA+by+Combining+Genetic+Alphabet+Expansion+Transcription+and+Copper-Free+Click+Chemistry.">Site-Specific Labeling of RNA by Combining Genetic Alphabet Expansion Transcription and Copper-Free Click Chemistry.</a> Nucl. Acids Res. 43, 6665-6676 (2015).</li><li>Lee, S. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mesoporous+Silica+Nanoparticle+Pretargeting+for+PET+Imaging+Based+on+a+Rapid+Bioorthogonal+Reaction+in+a+Living+Body.">Mesoporous Silica Nanoparticle Pretargeting for PET Imaging Based on a Rapid Bioorthogonal Reaction in a Living Body.</a> Angew. Chem. Int. Ed. 52, 10549-10552 (2013).</li><li>Sachin, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=F-18+Labeling+Protocol+of+Peptides+Based+on+Chemically+Orthogonal+Strain-Promoted+Cycloaddition+under+Physiologically+Friendly+Reaction+Conditions.">F-18 Labeling Protocol of Peptides Based on Chemically Orthogonal Strain-Promoted Cycloaddition under Physiologically Friendly Reaction Conditions.</a> Bioconjugate Chem. 23, 1680-1686 (2012).</li><li>Evans, H. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Copper-Free+Click+-+A+Promising+Tool+for+Pre-targeted+PET+Imaging.">Copper-Free Click - A Promising Tool for Pre-targeted PET Imaging.</a> Chem. Commun. 48, 991-993 (2012).</li><li>Campbell-Verduyn, L. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Strain-Promoted+Copper-Free+&amp;#34;Click&amp;#34;+Chemistry+for+18F+Radiolabeling+of+Bombesin.">Strain-Promoted Copper-Free &amp;#34;Click&amp;#34; Chemistry for 18F Radiolabeling of Bombesin.</a> Angew. Chem. Int. Ed. 50, 11117-11120 (2011).</li><li>Arumugam, S., Chin, J., Schirrmacher, R., Popik, V. V., Kostikov, A. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=18F]Azadibenzocyclooctyne+([18F]ADIBO):+A+Biocompatible+Radioactive+Labeling+Synthon+for+Peptides+using+Catalyst+Free+[3+2]+Cycloaddition.">18F]Azadibenzocyclooctyne ([18F]ADIBO): A Biocompatible Radioactive Labeling Synthon for Peptides using Catalyst Free [3+2] Cycloaddition.</a> Bioorg. Med. Chem. Lett. 21, 6987-6991 (2011).</li><li>Bouvet, V., Wuest, M., Wuest, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Copper-Free+Click+Chemistry+with+the+Short-Lived+Positron+Emitter+Fluorine-18.">Copper-Free Click Chemistry with the Short-Lived Positron Emitter Fluorine-18.</a> Org. Biomol. Chem. 9, 7393-7399 (2011).</li><li>Satpati, D., Bauer, N., Hausner, S. H., Sutcliffe, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthesis+of+[64Cu]DOTA-ADIBON3-Ala-PEG28-A20FMDV2+via+Copper-Free+Click+Chemistry+for+PET+Imaging+of+Integrin+&#945;v&#946;6.">Synthesis of [64Cu]DOTA-ADIBON3-Ala-PEG28-A20FMDV2 via Copper-Free Click Chemistry for PET Imaging of Integrin &#945;v&#946;6.</a> J. Radioanal. Nucl. Chem. 302, 765-771 (2014).</li><li>Lee, D. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Facile+Method+To+Radiolabel+Glycol+Chitosan+Nanoparticles+with+64Cu+via+Copper-Free+Click+Chemistry+for+MicroPET+Imaging.">Facile Method To Radiolabel Glycol Chitosan Nanoparticles with 64Cu via Copper-Free Click Chemistry for MicroPET Imaging.</a> Mol. Pharmaceutics. 10, 2190-2198 (2013).</li><li>Zeng, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=64Cu+Core-Labeled+Nanoparticles+with+High+Specific+Activity+via+Metal-Free+Click+Chemistry.">64Cu Core-Labeled Nanoparticles with High Specific Activity via Metal-Free Click Chemistry.</a> ACS Nano. 6, 5209-5219 (2012).</li><li>Jeon, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Radiosynthesis+and+in+vivo+Evaluation+of+[125I]2-4(iodophenethyl)-2-Methylmalonic+Acid+as+a+Potential+Radiotracer+for+Detection+of+Apoptosis.">Radiosynthesis and in vivo Evaluation of [125I]2-4(iodophenethyl)-2-Methylmalonic Acid as a Potential Radiotracer for Detection of Apoptosis.</a> J. Radioanal. Nucl. Chem. 308, 23-29 (2016).</li><li>Adam, M. J., Wilbur, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Radiohalogens+for+Imaging+and+Therapy.">Radiohalogens for Imaging and Therapy.</a> Chem. Soc. Rev. 34, 153-163 (2005).</li><li>Jeon, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Radiosynthesis+of+123I-Labeld+Hesperetin+for+Biodistribution+Study+of+Orally+Administered+Hesperetin.">Radiosynthesis of 123I-Labeld Hesperetin for Biodistribution Study of Orally Administered Hesperetin.</a> J. Radioanal. Nucl. Chem. 306, 437-443 (2015).</li><li>Kil, K. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+[123I]IPEB+and+[123I]IMPEB+as+SPECT+Radioligands+for+Metabotropic+Glutamate+Receptor+Subtype.">Development of [123I]IPEB and [123I]IMPEB as SPECT Radioligands for Metabotropic Glutamate Receptor Subtype.</a> ACS Med. Chem. Lett. 5, 652-656 (2014).</li><li>Chen, M. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Utility+of+I-123+Pretherapy+Scan+in+I-131+Radioiodine+Therapy+for+Thyroid+Cancer.">The Utility of I-123 Pretherapy Scan in I-131 Radioiodine Therapy for Thyroid Cancer.</a> Thyroid. 22, 304-309 (2012).</li><li>Jeon, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficient+Method+for+Iodine+Radioisotope+Labeling+of+Cyclooctyne-Containing+Molecules+using+Strain-Promoted+Copper-Free+Click+Reaction.">Efficient Method for Iodine Radioisotope Labeling of Cyclooctyne-Containing Molecules using Strain-Promoted Copper-Free Click Reaction.</a> Bioorg. Med. Chem. 23, 3303-3308 (2015).</li><li>Choi, M. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthesis+and+Evaluation+of+an+125I-Labeled+Azide+Prosthetic+Group+for+Efficient+and+Bioorthogonal+Radiolabeling+of+Cyclooctyne-Group+Containing+Molecules+using+Copper-Free+Click+Reaction.">Synthesis and Evaluation of an 125I-Labeled Azide Prosthetic Group for Efficient and Bioorthogonal Radiolabeling of Cyclooctyne-Group Containing Molecules using Copper-Free Click Reaction.</a> Bioorg. Med. Chem. Lett. 26, 875-878 (2016).</li><li>Kim, Y. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tumor+Targeting+and+Imaging+Using+Cyclic+RGD-PEGylated+Gold+Nanoparticle+Probes+with+Directly+Conjugated+Iodine-125.">Tumor Targeting and Imaging Using Cyclic RGD-PEGylated Gold Nanoparticle Probes with Directly Conjugated Iodine-125.</a> Small. 7, 2052-2060 (2011).</li></ol>

The authors have nothing to disclose.

באופן כללי, התשואה radiochemical הנצפה של אזיד מטוהרים 125 שכותרתו-I (1) היה 75 ± 10% (n = 8). Radiolabeling הושג עם 50-150 MBq של רדיואקטיביות, ותוצאות radiochemical הן די עקביות. אם [125 I] Nai (t 1/2 = 59.4 ד) כי עבר התפרקות רדיואקטיבית במשך יותר מחודש שמשה תגובת radioiodination, תשואת ra...

The strain-promoted copper-free click reaction between azides and cyclooctynes has been extensively applied to the efficient bioorthogonal labeling of a wide range of biomolecules, nanomaterials, and living subjects1-7. Due to the excellent site-specificity and rapid reaction rate of this conjugation reaction, it has also been used to synthesize radiolabeled tracers. A few 18F-labeled azide or DBCO prosthetic groups have been prepared for in vitro labeling of various cancers targeting peptides and antibodies, as well as for in vivo pre-targeted imaging of tumors8-13. In addition to these examples, the same conjugation reaction was applied to the metal-radioisotope-labeling of nanomaterials for positron emission tomography (PET) imaging studies14-16.
For several decades, radioactive iodines have been used for biomedical research and clinical trials through PET imaging (124I), single-photon emission computed tomography (SPECT) imaging (123I, 125I), and thyroid cancer treatment (131I)17-21. Therefore, an efficient method for radioactive iodine labeling is fundamentally important for various investigations, including molecular imaging studies, analysis of organ distribution of biomolecules, biomarker identification, and drug development. A copper-free click reaction strategy could be used in radioactive iodine labeling. However, this application has not been investigated as extensively as 18F-labeled biomolecules22-23. Here, we will provide a step-by-step protocol for the synthesis of an 125I-labeled azide for radiolabeling of DBCO-group-derived molecules. The procedures in the present report will include radioiodination of the stannylated precursor, purification steps with HPLC, and solid phase extraction. We also demonstrate efficient radiolabeling of DBCO-group-modified 13-nm-sized gold nanoparticles using the 125I-labeled azide. The detailed protocol in this report will help synthetic chemists understand a new radiolabeling methodology for the synthesis of radiolabeled products.

<table><tbody><tr><td>Chloramine T trihydrate</td><td>Sigma</td><td>402869</td><td></td></tr><tr><td>[&lt;sup&gt;125&lt;/sup&gt;I]NaI in aq. NaOH</td><td>Perkin-Elmer</td><td>NEZ033A010MC</td><td></td></tr><tr><td>Sodium metabisulfite&amp;nbsp;</td><td>Sigma</td><td>S9000</td><td></td></tr><tr><td>Formic acid</td><td>Sigma</td><td>251364</td><td></td></tr><tr><td>Sep-Pak tC18 plus cartridge</td><td>Waters</td><td>WAT036800</td><td></td></tr><tr><td>Dimethyl sulfoxide&amp;nbsp;</td><td>Sigma</td><td>D2650</td><td></td></tr><tr><td>Acetone</td><td>Sigma</td><td>650501</td><td></td></tr><tr><td>Ethanol</td><td>Sigma</td><td>459844</td><td></td></tr><tr><td>Gold(III) chloride trihydrate</td><td>Sigma</td><td>520918</td><td></td></tr><tr><td>Tween 20&amp;nbsp;</td><td>Sigma</td><td>P1379</td><td></td></tr><tr><td>DBCO PEG SH (MW 5,000)</td><td>NANOCS</td><td>PG2-DBTH-5k</td><td></td></tr><tr><td>TLC silica gel 60 F&lt;sub&gt;254&lt;/sub&gt;</td><td>Merck</td><td></td><td></td></tr><tr><td>Analytical HPLC</td><td>Agilent</td><td>1290 Infinity</td><td>Model number</td></tr><tr><td>Preparative HPLC</td><td>Agilent</td><td>1260 Infinity</td><td>Model number</td></tr><tr><td>Analytical C18 reverse-phase column</td><td>Agilent</td><td>Zorbax Eclipse XDB-C18</td><td></td></tr><tr><td>Preparative C18 reverse-phase column</td><td>Agilent</td><td>PrepHT XDB-C18</td><td></td></tr><tr><td>Radio TLC scanner</td><td>Bioscan</td><td>AR-2000</td><td>Model number</td></tr><tr><td>Radioisotope dose calibrator</td><td>Capintec, Inc</td><td>CRC -25R dose calibrator</td><td>Model number</td></tr></tbody></table>

an optimized protocol for the efficient radiolabeling of gold nanoparticles by using a 125i-labeled azide prosthetic group

Here, we demonstrate a detailed protocol for the radiosynthesis of a 125I-labeled azide prosthetic group and its application to the efficient radiolabeling of DBCO-group-functionalized gold nanoparticles using a copper-free click reaction. Radioiodination of the stannylated precursor (2) was carried out by using [125I]NaI and chloramine T as an oxidant at room temperature for 15 min. After HPLC purification of the crude product, the purified 125I-labeled azide (1) was obtained with high radiochemical yield (75 &plusmn; 10%, n = 8) and excellent radiochemical purity (&gt;99%). For the synthesis of radiolabeled 13-nm-sized gold nanoparticles, the DBCO-functionalized gold nanoparticles (3) were prepared by using a thiolated polyethylene glycol polymer. A copper-free click reaction between 1 and 3 gave the 125I-labeled gold nanoparticles (4) with more than 95% of radiochemical yield as determined by radio-thin-layer chromatography (radio-TLC). These results clearly indicate that the present radiolabeling method using a strain-promoted copper-free click reaction will be useful for the efficient and convenient radiolabeling of DBCO-group-containing nanomaterials.

תגובת radioiodination של מבשר stannylated (2) בוצעה באמצעות 150 MBq של [125 I] נאה, חומצה אצטית, ו chloramine T בטמפרטורת חדר למשך 15 דקות כדי לספק את המוצר רדיואקטיבי (1). לאחר HPLC טיהור preparative של התערובת הגולמית, המוצר הרצוי הושג עם 75 ± 10% (n = 8) של תשואת radioche...

Watch this Scientific Journal Video about An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group at JoVE.com

An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group

נוהל מפורט לסינתזה של אזיד 125 I-שכותרתו ואת radiolabeling של dibenzocyclooctyne (DBCO) -group מצומדות, 13 ננומטר בגודל ננו-חלקיקי זהב באמצעות תגובת לחץ ללא נחושת מתואר.

פרוטוקול אופטימיזציה עבור Radiolabeling היעיל של חלקיקי זהב על ידי שימוש<sup&gt; 125</sup&gt; שכותרתו לי אזיד תותבת קבוצה

Here, we demonstrate a detailed protocol for the radiosynthesis of a 125I-labeled azide prosthetic group and its application to the efficient ...

an-optimized-protocol-for-efficient-radiolabeling-gold-nanoparticles

Universidad San Sebastian

Research

JoVE Journal

Chemistry

An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group

8.1K Views.  Korea Atomic Energy Research Institute.  נוהל מפורט לסינתזה של אזיד 125 I-שכותרתו ואת radiolabeling של dibenzocyclooctyne (DBCO) -group מצומדות, 13 ננומטר בגודל ננו-חלקיקי זהב באמצעות תגובת לחץ ללא נחושת מתואר. 

Video: An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group

Efficient and Site-specific Antibody Labeling by Strain-promoted Azide-alkyne Cycloaddition

There are currently many chemical tools available to introduce chemical probes into proteins to study their structure and function. A useful method is protein conjugation by genetically introducing an unnatural amino acid containing a bioorthogonal functional group. This report describes a detailed protocol for site-specific antibody conjugation. The protocol includes experimental details for the genetic incorporation of an azide-containing amino acid, and the conjugation reaction by strain-promoted azide-alkyne cycloaddition (SPAAC). This strain-promoted reaction proceeds by simple mixing of the reacting molecules at physiological pH and temperature, and does not require additional reagents such as copper(I) ions and copper-chelating ligands. Therefore, this method would be useful for general protein conjugation and development of antibody drug conjugates (ADCs).

Here, we present a protocol to site-specifically introduce chemical probes into an antibody fragment by genetically incorporating an azide-containing amino acid, and subsequently coupling the azide with a chemical probe by strain-promoted azide-alkyne cycloaddition (SPAAC).

יעיל אתר ספציפי תיוג נוגדן ידי cycloaddition יזיד-אלקין קדם-Strain

There are currently many chemical tools available to introduce chemical probes into proteins to study their structure and function. A useful method is ...

Biochemistry

The Bioconjugation and Radiosynthesis of 89Zr-DFO-labeled Antibodies

The exceptional affinity, specificity, and selectivity of antibodies make them extraordinarily attractive vectors for tumor-targeted PET radiopharmaceuticals. Due to their multi-day biological half-life, antibodies must be labeled with positron-emitting radionuclides with relatively long physical decay half-lives. Traditionally, the positron-emitting isotopes 124I (t1/2 = 4.18 d), 86Y (t1/2 = 14.7 hr), and 64Cu (t1/2 = 12.7 hr) have been used to label antibodies for PET imaging. More recently, however, the field has witnessed a dramatic increase in the use of the positron-emitting radiometal 89Zr in antibody-based PET imaging agents. 89Zr is a nearly ideal radioisotope for PET imaging with immunoconjugates, as it possesses a physical half-life (t1/2 = 78.4 hr) that is compatible with the in vivo pharmacokinetics of antibodies and emits a relatively low energy positron that produces high resolution images. Furthermore, antibodies can be straightforwardly labeled with 89Zr using the siderophore-derived chelator desferrioxamine (DFO). In this protocol, the prostate-specific membrane antigen targeting antibody J591 will be used as a model system to illustrate (1) the bioconjugation of the bifunctional chelator DFO-isothiocyanate to an antibody, (2) the radiosynthesis and purification of a 89Zr-DFO-mAb radioimmunoconjugate, and (3) in vivo PET imaging with an 89Zr-DFO-mAb radioimmunoconjugate in a murine model of cancer.

The Bioconjugation and Radiosynthesis of 89Zr-DFO-labeled Antibodies

Due to its multi-day radioactive half-life and favorable decay properties, the positron-emitting radiometal 89Zr is extremely well-suited for use in antibody-based radiopharmaceuticals for PET imaging. In this protocol, the bioconjugation, radiosynthesis, and preclinical application of 89Zr-labeled antibodies will be described.

Bioconjugation וRadiosynthesis של<sup&gt; 89</sup&gt; נוגדנים שכותרתו ZR-DFO

The exceptional affinity, specificity, and selectivity of antibodies make them extraordinarily attractive vectors for tumor-targeted PET ...

Harnessing the Bioorthogonal Inverse Electron Demand Diels-Alder Cycloaddition for Pretargeted PET Imaging

Due to their exquisite affinity and specificity, antibodies have become extremely promising vectors for the delivery of radioisotopes to cancer cells for PET imaging. However, the necessity of labeling antibodies with radionuclides with long physical half-lives often results in high background radiation dose rates to non-target tissues. In order to circumvent this issue, we have employed a pretargeted PET imaging strategy based on the inverse electron demand Diels-Alder cycloaddition reaction. The methodology decouples the antibody from the radioactivity and thus exploits the positive characteristics of antibodies, while eschewing their pharmacokinetic drawbacks. The system is composed of four steps: (1) the injection of a mAb-trans-cyclooctene (TCO) conjugate; (2) a localization time period during which the antibody accumulates in the tumor and clears from the blood; (3) the injection of the radiolabeled tetrazine; and (4) the in vivo click ligation of the components followed by the clearance of excess radioligand. In the example presented in the work at hand, a 64Cu-NOTA-labeled tetrazine radioligand and a trans-cyclooctene-conjugated humanized antibody (huA33) were successfully used to delineate SW1222 colorectal cancer tumors with high tumor-to-background contrast. Further, the pretargeting methodology produces high quality images at only a fraction of the radiation dose to non-target tissue created by radioimmunoconjugates directly labeled with 64Cu or 89Zr. Ultimately, the modularity of this protocol is one of its greatest assets, as the trans-cyclooctene moiety can be appended to any non-internalizing antibody, and the tetrazine can be attached to a wide variety of radioisotopes.

The bioorthogonal inverse electron demand Diels-Alder cycloaddition has been harnessed to create an effective and modular pretargeted PET imaging strategy for cancer. In this protocol, the steps of this methodology are described in the context of a model system employing the colorectal cancer targeted antibody huA33 and a 64Cu-labeled radioligand.

רתימת ביקוש Bioorthogonal הפוך אלקטרונים דילס-Alder cycloaddition לPretargeted PET הדמיה

Due to their exquisite affinity and specificity, antibodies have become extremely promising vectors for the delivery of radioisotopes to cancer cells for ...

Bioengineering

Pretargeted Radioimmunotherapy Based on the Inverse Electron Demand Diels-Alder Reaction

While radioimmunotherapy (RIT) is a promising approach for the treatment of cancer, the long pharmacokinetic half-life of radiolabeled antibodies can result in high radiation doses to healthy tissues. Perhaps not surprisingly, several different strategies have been developed to circumvent this troubling limitation. One of the most promising of these approaches is pretargeted radioimmunotherapy (PRIT). PRIT is predicated on decoupling the radionuclide from the immunoglobulin, injecting them separately, and then allowing them to combine in vivo at the target tissue. This approach harnesses the exceptional tumor-targeting properties of antibodies while skirting their pharmacokinetic drawbacks, thereby lowering radiation doses to non-target tissues and facilitating the use of radionuclides with half-lives that are considered too short for use in traditional radioimmunoconjugates. Over the last five years, our laboratory and others have developed an approach to in vivo pretargeting based on the inverse electron-demand Diels-Alder (IEDDA) reaction between trans-cyclooctene (TCO) and tetrazine (Tz). This strategy has been successfully applied to pretargeted positron emission tomography (PET) and single-photon emission computed tomography (SPECT) imaging with a variety of antibody-antigen systems. In a pair of recent publications, we have demonstrated the efficacy of IEDDA-based PRIT in murine models of pancreatic ductal adenocarcinoma and colorectal carcinoma. In this protocol, we describe protocols for PRIT using a 177Lu-DOTA-labeled tetrazine radioligand ([177Lu]Lu-DOTA-PEG7-Tz) and a TCO-modified variant of the colorectal cancer targeting huA33 antibody (huA33-TCO). More specifically, we will describe the construction of huA33-TCO, the synthesis and radiolabeling of [177Lu]Lu-DOTA-PEG7-Tz, and the performance of in vivo biodistribution and longitudinal therapy studies in murine models of colorectal carcinoma.

This protocol describes the synthesis and characterization of a trans-cyclooctene (TCO)-modified antibody and a 177Lu-labeled tetrazine (Tz) radioligand for pretargeted radioimmunotherapy (PRIT). In addition, it details the use of these two constructs for in vivo biodistribution and longitudinal therapy studies in a murine model of colorectal cancer.

Radioimmunotherapy pretargeted התגובה תגובת דילס-אלדר דרישה אלקטרון הופכי

While radioimmunotherapy (RIT) is a promising approach for the treatment of cancer, the long pharmacokinetic half-life of radiolabeled antibodies can ...

Development of a 68Gallium-Labeled D-Peptide PET Tracer for Imaging Programmed Death-Ligand 1 Expression

The development of immune checkpoint blockade therapy based on programmed cell death-protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) has revolutionized cancer therapies in recent years. However, only a fraction of patients responds to PD-1/PD-L1 inhibitors, owing to the heterogeneous expression of PD-L1 in tumor cells. This heterogeneity presents a challenge in the precise detection of tumor cells by the commonly used immunohistochemistry (IHC) approach. This situation calls for better methods to stratify patients who will benefit from immune checkpoint blockade therapy, to improve treatment efficacy. Positron emission tomography (PET) enables real-time visualization of the whole-body PD-L1 expression in a noninvasive way. Therefore, there is a need for the development of radiolabeled tracers to detect PD-L1 distribution in tumors through PET imaging.
Compared to their L-counterparts, dextrorotary (D)-peptides have properties such as proteolytic resistance and remarkably prolonged metabolic half-lives. This study designed a new method to detect PD-L1 expression based on PET imaging of 68Ga-labeled PD-L1-targeted D-peptide, a D-dodecapeptide antagonist (DPA), in tumor-bearing mice. The results showed that the [68Ga]DPA can specifically bind to PD-L1-overexpressing tumors in vivo, and showed favorable stability as well as excellent imaging ability, suggesting that [68Ga]DPA-PET is a promising approach for the assessment of PD-L1 status in tumors.

Development of a 68Gallium-Labeled D-Peptide PET Tracer for Imaging Programmed Death-Ligand 1 Expression

This study developed a noninvasive and real-time method to evaluate the distribution of programmed death-ligand 1 in the whole body, based on positron emission tomographic imaging of [68Ga] D-dodecapeptide antagonist. This technique has advantages over conventional immunohistochemistry and improves the efficiency of identifying appropriate patients who will benefit from immune checkpoint blockade therapy.

פיתוח של 68גלייום מסומן D-פפטיד PET Tracer עבור הדמיה מתוכנת מוות-ליגנד 1 ביטוי

The development of immune checkpoint blockade therapy based on programmed cell death-protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) has revolutionized ...

Cancer Research

Positron Emission Tomography Imaging of Cell Trafficking: A Method of Cell Radiolabeling

Stem cell and chimeric antigen receptor (CAR) T-cell therapies are emerging as promising therapeutics for organ regeneration and as immunotherapy for various cancers. Despite significant progress having been made in these areas, there is still more to be learned to better understand the pharmacokinetics and pharmacodynamics of the administered therapeutic cells in the living system. For noninvasive, in vivo tracking of cells with positron emission tomography (PET), a novel [89Zr]Zr-p-isothiocyanatobenzyl-desferrioxamine ([89Zr]Zr-DBN)-mediated cell radiolabeling method has been developed utilizing 89Zr (t1/2 78.4 h). The present protocol describes a [89Zr]Zr-DBN-mediated, ready-to-use, radiolabeling synthon for direct radiolabeling of variety of cells, including mesenchymal stem cells, lineage-guided cardiopoietic stem cells, liver regenerating hepatocytes, white blood cells, melanoma cells, and dendritic cells. The developed methodology enables noninvasive PET imaging of cell trafficking for up to 7 days post-administration without affecting the nature or the function of the radiolabeled cells. Additionally, this protocol describes a stepwise method for the radiosynthesis of [89Zr]Zr-DBN, biocompatible formulation of [89Zr]Zr-DBN, preparation of cells for radiolabeling, and finally the radiolabeling of cells with [89Zr]Zr-DBN, including all the intricate details needed for the successful radiolabeling of cells.

Presented here is a protocol to radiolabel cells with a positron emission tomography (PET) radioisotope, 89Zr (t1/2 78.4 h), using a ready-to-use radiolabeling synthon, [89Zr]Zr-p-isothiocyanatobenzyl-desferrioxamine ([89Zr]Zr-DBN). Radiolabeling cells with [89Zr]Zr-DBN allows noninvasive tracking and imaging of administered radiolabeled cells in the body with PET for up to 7 days post-administration.

טומוגרפיה של פליטת פוזיטרונים הדמיה של סחר בתאים: שיטה של תיוג רדיו תאים

Stem cell and chimeric antigen receptor (CAR) T-cell therapies are emerging as promising therapeutics for organ regeneration and as immunotherapy for ...

Radiosynthesis and PET/CT Imaging of 68Ga-Labelled Nanobody in Xenograft Models

Radiosynthesis, Quality Control, and Small Animal Positron Emission Tomography Imaging of 68Ga-Labelled Nano Molecules

Small animal Positron Emission Tomography (PET) and Single Photon Emission Computed Tomography (SPECT) imaging techniques are crucial in preclinical cancer research, necessitating meticulous attention to radiotracer synthesis, quality assurance, and in vivo injection protocols. This study presents a comprehensive workflow tailored to enhance the robustness and reproducibility of small animal PET experiments. The synthesis process in the radiochemistry laboratory using 68Ga is detailed, highlighting stringent quality control and assurance protocols for each radiotracer production. Parameters such as concentration, molar activity, pH, and purity are rigorously monitored, aligning with standards applicable to human studies. This methodology introduces streamlined syringe preparation and a custom-designed 30G cannula for precise intravenous injections into mice. Monitoring of animal health during scanning, including temperature and heart rate, ensures their well-being throughout the procedure. Dosages for PET and SPECT scans are predetermined to balance data acquisition with minimizing radiation exposure to animals and researchers. Similarly, CT scans employ pre-programmed settings to limit radiation exposure, especially pertinent in long-term studies assessing treatment effects. By optimizing these steps, the workflow aims to standardize procedures, reduce variability, and enhance the quality of small animal PET/SPECT/CT imaging. This resource provides valuable insights for researchers seeking to improve the accuracy and reliability of preclinical investigations in molecular imaging, ultimately advancing the field.

This paper describes the radiosynthesis, formulation, quality control of a new radiolabeled probe (i.e., 68Ga-labeled nanobody NM-02), and its use for small animal PET/CT imaging in a xenograft model.

Small animal Positron Emission Tomography (PET) and Single Photon Emission Computed Tomography (SPECT) imaging techniques are crucial in preclinical ...

Murine Lymphocyte Labeling by 64Cu-Antibody Receptor Targeting for In Vivo Cell Trafficking by PET/CT

This protocol illustrates the production of 64Cu and the chelator conjugation/radiolabeling of a monoclonal antibody (mAb) followed by murine lymphocyte cell culture and 64Cu-antibody receptor targeting of the cells. In vitro evaluation of the radiolabel and non-invasive in vivo cell tracking in an animal model of an airway delayed-type hypersensitivity reaction (DTHR) by PET/CT are described.
In detail, the conjugation of a mAb with the chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) is shown. Following the production of radioactive 64Cu, radiolabeling of the DOTA-conjugated mAb is described. Next, the expansion of chicken ovalbumin (cOVA)-specific CD4+ interferon (IFN)-&#947;-producing T helper cells (cOVA-TH1) and the subsequent radiolabeling of the cOVA-TH1 cells are depicted. Various in vitro techniques are presented to evaluate the effects of 64Cu-radiolabeling on the cells, such as the determination of cell viability by trypan blue exclusion, the staining for apoptosis with Annexin V for flow cytometry, and the assessment of functionality by IFN-&#947; enzyme-linked immunosorbent assay (ELISA). Furthermore, the determination of the radioactive uptake into the cells and the labeling stability are described in detail. This protocol further describes how to perform cell tracking studies in an animal model for an airway DTHR and, therefore, the induction of cOVA-induced acute airway DHTR in BALB/c mice is included. Finally, a robust PET/CT workflow including image acquisition, reconstruction, and analysis is presented.
The 64Cu-antibody receptor targeting approach with subsequent receptor internalization provides high specificity and stability, reduced cellular toxicity, and low efflux rates compared to common PET-tracers for cell labeling, e.g.64Cu-pyruvaldehyde bis(N4-methylthiosemicarbazone) (64Cu-PTSM). Finally, our approach enables non-invasive in vivo cell tracking by PET/CT with an optimal signal-to-background ratio for 48 h. This experimental approach can be transferred to different animal models and cell types with membrane-bound receptors that are internalized.

Murine Lymphocyte Labeling by 64Cu-Antibody Receptor Targeting for In Vivo  Cell Trafficking by PET/CT

Following the preparation of a 64Cu-modified monoclonal antibody binding to a transgenic murine T cell receptor, T cells are radiolabeled in vivo, analyzed for viability, functionality, labeling stability and apoptosis, and adoptively transferred into mice with an airway delayed-type hypersensitivity reaction for non-invasive imaging by positron emission tomography/computed tomography (PET/CT).

Murine לימפוציטים תיוג ידי<sup&gt; 64</sup&gt; Cu-נוגדן קולטן מיקוד עבור<em&gt; In vivo</em&gt; סחר Cell על ידי PET / CT

This protocol illustrates the production of 64Cu and the chelator conjugation/radiolabeling of a monoclonal antibody (mAb) followed by murine lymphocyte ...

Immunology and Infection

<meta charset="utf8"></head><body>סינתזה יעילה של רדיוטרייסרים פחמן-11 באמצעות מתילציה בשיטת לולאה

Enhancing Efficiency and Radiolabeling Yields of Carbon-11 Radioligands for Clinical Research Using the Loop Method

A successful positron emission tomography imaging program involving carbon-11 radiotracers demands fast, efficient, and reliable synthesis methods, requiring an on-site cyclotron and radiochemistry group, as well as clinical staff trained to operate under the unique constraints of the carbon-11 radionuclide. This study examines the merits and advantages of a captive solvent &#39;loop method&#39; of radiolabeling four tracers with the carbon-11 radionuclide, producing the radioligands [11C]ER-176, [11C]MRB, [11C]mHED, and [11C]PiB.
The &#39;loop method&#39; is compared against the traditional reactor-based method of carbon-11 methylation in the course of synthesizing the same radiotracers on the identical automated platform. Further, a complete overview of the clinical research preparation of the [11C]ER-176 radiotracer is presented. As demonstrated by the production of [11C]ER-176, the captive solvent &#39;loop method&#39; of heterogeneous alkylation proved to be more efficient, with excellent radiochemical purity (99.6 &#177; 0.6%, n = 25), higher and more consistent radiochemical yield (end of synthesis (EOS) = 5.4 &#177; 2.2 GBq, n = 25) compared to the reactor method (EOS = 1.6 &#177; 0.5 GBq, n = 6), increased molar activity (loop method = 194 &#177; 66 GBq/&#181;mol, n = 25; reactor method = 132 &#177; 78 GBq/&#181;mol, n = 6), along with an average 5 min shorter reaction sequence.

<meta charset="utf8"></head><body>שיפור היעילות ותפוקות התיוג הרדיואקטיבי של רדיוליגנדים פחמן-11 למחקר קליני בשיטת לולאה

This protocol presents a detailed procedure to prepare four carbon-11 radiotracers by way of the &#8220;loop method&#8221; [11C]methylation. We describe the procedure to synthesize [11C]ER-176, &#160;with data for three additional radiotracers: [11C]MRB, [11C]mHED, and [11C]PiB. The loop method provides an efficient synthesis with increased radiochemical yields against traditional reaction vessel [11C]methylation.

שיפור היעילות ותפוקות התיוג הרדיואקטיבי של רדיוליגנדים פחמן-11 למחקר קליני בשיטת לולאה

A successful positron emission tomography imaging program involving carbon-11 radiotracers demands fast, efficient, and reliable synthesis methods, ...

פרוטוקול אופטימיזציה עבור Radiolabeling היעיל של חלקיקי זהב על ידי שימוש

Summary

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שיפור היעילות ותפוקות התיוג הרדיואקטיבי של רדיוליגנדים פחמן-11 למחקר קליני בשיטת לולאה

שיפור היעילות ותפוקות התיוג הרדיואקטיבי של רדיוליגנדים פחמן-11 למחקר קליני בשיטת לולאה