التفاصيل البيبلوغرافية
| العنوان: |
Quantification of In Vivo Target Engagement Using Microfluidic Activity-Based Protein Profiling. |
| المؤلفون: |
Reardon HT; Abide Therapeutics, San Diego, CA, USA., Herbst RA; Abide Therapeutics, San Diego, CA, USA., Henry CL; Abide Therapeutics, San Diego, CA, USA., Herbst DM; Abide Therapeutics, San Diego, CA, USA., Ngo N; Abide Therapeutics, San Diego, CA, USA., Cisar JS; Abide Therapeutics, San Diego, CA, USA.; Janssen Research & Development, Spring House, PA., Weber OD; Abide Therapeutics, San Diego, CA, USA., Niphakis MJ; Abide Therapeutics, San Diego, CA, USA., O'Neill GP; Abide Therapeutics, San Diego, CA, USA. |
| المصدر: |
SLAS technology [SLAS Technol] 2019 Oct; Vol. 24 (5), pp. 489-498. Date of Electronic Publication: 2019 Jun 14. |
| نوع المنشور: |
Journal Article |
| اللغة: |
English |
| بيانات الدورية: |
Publisher: SAGE Publications Country of Publication: United States NLM ID: 101697564 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 2472-6311 (Electronic) Linking ISSN: 24726303 NLM ISO Abbreviation: SLAS Technol Subsets: MEDLINE |
| أسماء مطبوعة: |
Original Publication: Thousand Oaks, CA : SAGE Publications, [2017]- |
| مواضيع طبية MeSH: |
Microfluidics/*methods , Proteomics/*methods, Algorithms ; Animals ; Biological Assay ; Mice ; Molecular Weight ; Reproducibility of Results |
| مستخلص: |
Accurate measurement of drug-target interactions in vivo is critical for both preclinical development and translation to clinical studies, yet many assays rely on indirect measures such as biomarkers associated with target activity. Activity-based protein profiling (ABPP) is a direct method of quantifying enzyme activity using active site-targeted small-molecule covalent probes that selectively label active but not inhibitor-bound enzymes. Probe-labeled enzymes in complex proteomes are separated by polyacrylamide gel electrophoresis and quantified by fluorescence imaging. To accelerate workflows and avoid imaging artifacts that make conventional gels challenging to quantify, we adapted protocols for a commercial LabChip GXII microfluidic instrument to permit electrophoretic separation of probe-labeled proteins in tissue lysates and plasma, and quantification of fluorescence (probe/protein labeling ratio of 1:1). Electrophoretic separation on chips occurred in 40 s per sample, and instrument software automatically identified and quantified peaks, resulting in an overall time savings of 3-5 h per 96-well sample plate. Calculated percent inhibition was not significantly different between the two formats. Chip performance was consistent between chips and sample replicates. Conventional gel imaging was more sensitive but required five times higher sample volume than microfluidic chips. Microfluidic chips produced results comparable to those of gels but with much lower sample consumption, facilitating assay miniaturization for scarce biological samples. The time savings afforded by microfluidic electrophoresis and automatic quantification has allowed us to incorporate microfluidic ABPP early in the drug discovery workflow, enabling routine assessments of tissue distribution and engagement of targets and off-targets in vivo. |
| فهرسة مساهمة: |
Keywords: ABPP; LabChip; MGLL; monoacylglycerol lipase; protein chip |
| تواريخ الأحداث: |
Date Created: 20190615 Date Completed: 20200417 Latest Revision: 20200417 |
| رمز التحديث: |
20221213 |
| DOI: |
10.1177/2472630319852303 |
| PMID: |
31199699 |
| قاعدة البيانات: |
MEDLINE |
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