کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8416893 | 1545655 | 2018 | 5 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
FACS isolation of live mouse eosinophils at high purity via a protocol that does not target Siglec F
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کلمات کلیدی
FACSGUSBITIMAspergillus fumigatus - آسپرژیلوس فومیگاتوسAllergen - آلرژنinflammation - التهاب( توروم) Interleukin-5 - اینترلوکین -5beta-glucuronidase - بتا گلوکورونیدازfluorescence-activated cell sorter - فلورسانس فعال سلول مرتب سازImmunoreceptor tyrosine-based inhibitory motif - مویر مهارکننده مبتنی بر تیروزین ایمونورزپتور
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
بیوتکنولوژی یا زیستفناوری
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چکیده انگلیسی
Flow cytometry protocols designed to identify mouse eosinophils typically target Siglec F, an α-2,3-sialic acid binding transmembrane protein expressed universally on cells of this lineage. While a convenient target, antibody-mediated ligation of Siglec F induces eosinophil apoptosis, which limits its usefulness for isolations that are to be followed by functional and/or gene expression studies. We present here a method for FACS isolation which does not target Siglec F and likewise utilizes no antibodies targeting IL5Rα (CD125) or CCR3. Single cell suspensions are prepared from lungs of mice that were sensitized and challenged with Aspergillus fumigatus antigens; eosinophils were identified and isolated by FACS as live SSChi/FSChi CD11câ Gr1â/loMHCIIâ cells. This strategy was also effective for eosinophil isolation from the lungs of IL5tg mice. Purity by visual inspection of stained cytospin preparations and by Siglec F-diagnostic flow cytometry was 98-99% and 97-99%, respectively. Eosinophils isolated by this method (yield, ~ 4 Ã 106/mouse) generated high-quality RNA suitable for gene expression analysis.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Immunological Methods - Volume 454, March 2018, Pages 27-31
Journal: Journal of Immunological Methods - Volume 454, March 2018, Pages 27-31
نویسندگان
Wendy E. Geslewitz, Caroline M. Percopo, Helene F. Rosenberg,