Thu, 21 Aug 2008 15:10:37 BST CiteULike: jyuh Benya http://www.citeulike.org/user/jyuh/author/Benya CiteULike.org en-gb Copyright © 2004-2008 citeulike.org http://www.citeulike.org/user/jyuh/article/965051 <i>J Histochem Cytochem, Vol. 48, No. 2. (February 2000), pp. 303-312.</i><br /><br />Currently available techniques for performing quantitative immunohistochemistry (Q-IHC) rely upon pixel-counting algorithms and therefore cannot provide information as to the absolute amount of chromogen present. We describe a novel algorithm for true Q-IHC based on calculating the cumulative signal strength, or energy, of the digital file representing any portion of an image. This algorithm involves subtracting the energy of the digital file encoding the control image (i.e., not exposed to antibody) from that of the experimental image (i.e., antibody-treated). In this manner, the absolute amount of antibody-specific chromogen per pixel can be determined for any cellular region or structure. (J Histochem Cytochem 48:303-311, 2000) Quantitative immunohistochemistry by measuring cumulative signal strength using commercially available software photoshop and matlab. KA Matkowskyj D Schonfeld RV Benya J Histochem Cytochem, Vol. 48, No. 2. (February 2000), pp. 303-312. 2006-11-28T08:46:22-00:00 2000 J Histochem Cytochem 0022-1554 48 2 303 312 imaging method http://www.citeulike.org/user/jyuh/article/2772625 <i>The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society, Vol. 51, No. 2. (February 2003), pp. 205-214.</i><br /><br />We previously demonstrated that quantitative immunohistochemistry (Q-IHC) performed by measuring the cumulative signal strength of the digital file encoding an image can be used to determine the absolute amount of chromogen present per pixel. We now show that Q-IHC so performed can be used to accurately determine the amount of peptide hormone receptor of interest in archived tissues. To do this we transfected Balb 3T3 fibroblasts with the cDNA encoding the human receptor for gastrin-releasing peptide (GRP), and selected six cell lines stably expressing between 10(2) and 10(6) receptors/cell. These cell lines were fixed in formalin, embedded in paraffin, and treated with antipeptide antibodies against the GRP receptor, followed by DAB chromogen to identify bound antibody. Images were acquired using a 4.9 million pixel digital scanning 24-bit RGB camera, saved in TIFF format, and used for subsequent analysis. Q-IHC was performed after digitally dissecting out the relevant portion of the image for analysis, and processing using a program written in C (available at http://www.uic.edu/com/dom/gastro/Freedownloads.html). Under the conditions defined here, chromogen quantity as determined by Q-IHC tightly correlated with GRP receptor number (r(2)=0.867) in these cell lines. Using the conversion factor identified as a result of these studies, we then determined GRP receptor number on eight randomly selected, archived human colon cancers. Overall GRP receptor expression in colon cancer depended on the degree to which cells within any particular tumor were differentiated, with well-differentiated cells expressing the greatest numbers of receptors (approximately 55,000 +/- 10,000 sites/cell). These studies indicate that Q-IHC can be used to determine receptor quantity in archived tissues and other samples of limited quantity. Quantitative immunohistochemistry by measuring cumulative signal strength accurately measures receptor number. KA Matkowskyj R Cox RT Jensen RV Benya The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society, Vol. 51, No. 2. (February 2003), pp. 205-214. 2008-05-08T15:39:34-00:00 2003 The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 0022-1554 51 2 205 214 no-tag