creators_name: Kaur, R
creators_name: Dikshit, Kanak L
creators_name: Raje, Manoj
type: article
datestamp: 2012-01-05 15:11:11
lastmod: 2012-03-29 09:48:02
metadata_visibility: show
title: Pre-screening for antigen detectability in cells: a TEM-based solid phase digital immunogold detection method utilizing ultra low volumes of reagents.
ispublished: pub
subjects: QR
full_text_status: restricted
keywords: Dot blot, immunodetection, immunogold labelling, polystyrene films,
pre-screening, solid phase, transmission electron microscopy.
note: Copyright of this article belongs to Wiley.
abstract: Rapid and sensitive pre-screening for the presence of antigens in cell samples and confirmation of reactivity of antibodies, before proceeding with electron microscopy, is highly desirable. Most of the methods developed for this purpose are generally not very efficient and suitable for dealing with very small volumes of sample and reagents. In this work we present a simple, sensitive and rapid solid phase transmission electron microscope (TEM) based method for the detection of picogram (pg) levels of soluble antigens using as little as 10 micro L of reagents. Protein was adsorbed onto grids coated with polystyrene films to form the solid phase. The presence of antigen was detected using immunogold labelling. Gold particles adhering to the film were visualized and counted in a TEM providing a digital signal. This method was 100-fold more sensitive than dot blot in detection of rabbit IgG. We have demonstrated the utility of this technique by screening for Vitreoscilla haemoglobin (VHb) antigen in cell lysates and confirming the results directly with immunogold labelling transmission electron microscopy of cell sections.
date: 2002-11
date_type: published
publication: Journal of microscopy
volume: 208
number: Pt 2
publisher: Wiley
pagerange: 100-7
refereed: TRUE
issn: 0022-2720
official_url: http://onlinelibrary.wiley.com/doi/10.1046/j.1365-2818.2002.01072.x/pdf
related_url_url: http://onlinelibrary.wiley.com/doi/10.1046/j.1365-2818.2002.01072.x/pdf
related_url_type: pub
citation:   Kaur, R and Dikshit, Kanak L and Raje, Manoj  (2002) Pre-screening for antigen detectability in cells: a TEM-based solid phase digital immunogold detection method utilizing ultra low volumes of reagents.  Journal of microscopy, 208 (Pt 2).  pp. 100-7.  ISSN 0022-2720     
document_url: http://crdd.osdd.net/open/259/1/raje2002.2.pdf