<html><body><div style="color:#000; background-color:#fff; font-family:verdana, helvetica, sans-serif;font-size:10pt"><div style="RIGHT: auto"><SPAN style="RIGHT: auto">Ya Peter, I know what yo<VAR id=yui-ie-cursor></VAR>u are saying about studying the expression profile of the transcriptome, one has to be careful regarding contaminating RNA from macrophages, in that case yes, one has to depend on axenic amastigotes which are much better purity wise as well as number wise, Thanks</SPAN></div>
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<DIV style="RIGHT: auto"><I style="RIGHT: auto"><FONT style="RIGHT: auto" color=#0000bf>sunil</FONT></I></DIV>
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<DIV style="BORDER-BOTTOM: #ccc 1px solid; BORDER-LEFT: #ccc 1px solid; PADDING-BOTTOM: 0px; LINE-HEIGHT: 0; MARGIN: 5px 0px; PADDING-LEFT: 0px; PADDING-RIGHT: 0px; HEIGHT: 0px; FONT-SIZE: 0px; BORDER-TOP: #ccc 1px solid; BORDER-RIGHT: #ccc 1px solid; PADDING-TOP: 0px" class=hr readonly="true" contenteditable="false"></DIV><B><SPAN style="FONT-WEIGHT: bold">From:</SPAN></B> Peter Myler <peter.myler@seattlebiomed.org><BR><B><SPAN style="FONT-WEIGHT: bold">To:</SPAN></B> 'Sunil Arora' <skarora_in@yahoo.com>; Clos Joachim <clos@bni-hamburg.de>; Leish-L email post <Leish-l@lineu.icb.usp.br> <BR><B><SPAN style="FONT-WEIGHT: bold">Cc:</SPAN></B> "danz@bi.technion.ac.il" <danz@bi.technion.ac.il> <BR><B><SPAN style="FONT-WEIGHT: bold">Sent:</SPAN></B> Saturday, 30 March 2013 1:39 AM<BR><B><SPAN style="FONT-WEIGHT: bold">Subject:</SPAN></B> RE: [Leish-l] Fwd: hot spots<BR></FONT></DIV><BR>
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<div class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Calibri', 'sans-serif'; COLOR: #1f497d; FONT-SIZE: 11pt">Sunil:</SPAN></div>
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<div style="RIGHT: auto" class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Calibri', 'sans-serif'; COLOR: #1f497d; FONT-SIZE: 11pt">Axenic amastigotes (Ax) are much more than “stressed-out promastigotes”. We (and others) have shown they are very similar (although not identical) to macrophage-derived amastigotes (Am) in terms of global mRNA and protein expression levels. While it would obviously be better to work with Am, it is often technically difficult due to low numbers and contamination with macrophage RNA and protein. In addition, it is my contention that the time taken to isolate Am from the macrophages makes them “stressed-out amastigotes”, which are less physiological relevant than Ax. We need to approach this discussion from a position of knowledge and data (i.e. understanding exactly which genes are expressed differently in Am and Ax) than belief. Hopefully, we will be able to publish the results of
our RNA-seq studies comparing promastigotes, axenic amastigotes and macrophage-derived amastigotes (without purification) in the not too distant future.</SPAN></div>
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<div class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Calibri', 'sans-serif'; COLOR: #1f497d; FONT-SIZE: 11pt">Peter </SPAN></div>
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<div class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: navy; FONT-SIZE: 10pt">Peter J. Myler, Ph.D.</SPAN><SPAN style="COLOR: navy"></SPAN></div>
<div class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: navy; FONT-SIZE: 10pt">Professor</SPAN><SPAN style="COLOR: navy"><BR></SPAN><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: navy; FONT-SIZE: 10pt">Seattle BioMed<BR>307 Westlake Ave N., Suite 500<BR>Seattle, WA 98109-5219<BR>USA</SPAN><SPAN style="COLOR: navy"> <BR></SPAN><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: navy; FONT-SIZE: 10pt">(206) 256-7332 (office)</SPAN><SPAN style="COLOR: navy"><BR></SPAN><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: navy; FONT-SIZE: 10pt">(206) 256-7229 (FAX)</SPAN><SPAN style="COLOR: navy"> <BR></SPAN><SPAN style="COLOR: #1f497d"><A href="mailto:peter.myler@sbri.org" rel=nofollow target=_blank ymailto="mailto:peter.myler@sbri.org">peter.myler@sbri.org</A></SPAN></div>
<div class=yiv1947419759MsoNormal><SPAN style="COLOR: navy">Affiliate Professor <BR>Departments of Global Health &<BR>Biomedical Informatics and Medical Education <BR>University of Washington <BR>Seattle, WA 98195 <BR>Box 357238 <BR></SPAN><SPAN style="COLOR: #1f497d"><A href="mailto:mylerpj@u.washington.edu" rel=nofollow target=_blank ymailto="mailto:mylerpj@u.washington.edu">mylerpj@u.washington.edu</A></SPAN><SPAN style="COLOR: navy"> </SPAN></div></DIV>
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<div class=yiv1947419759MsoNormal><B><SPAN style="FONT-FAMILY: 'Tahoma', 'sans-serif'; FONT-SIZE: 10pt">From:</SPAN></B><SPAN style="FONT-FAMILY: 'Tahoma', 'sans-serif'; FONT-SIZE: 10pt"> leish-l-bounces@lineu.icb.usp.br [mailto:leish-l-bounces@lineu.icb.usp.br] <B>On Behalf Of </B>Sunil Arora<BR><B>Sent:</B> Monday, March 11, 2013 8:42 AM<BR><B>To:</B> Clos Joachim; Leish-L email post<BR><B>Subject:</B> Re: [Leish-l] Fwd: hot spots</SPAN></div></DIV></DIV>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: black; FONT-SIZE: 10pt">I agree...axenic amastigotes are more like stressed out promastigotes... I always prefer working with something inside the macrophages as a model for testing drugs or immunomodulators which I believe are closer to actual intracellular amastigotes in the infected host</SPAN></div></DIV>
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<div class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: black; FONT-SIZE: 10pt">best</SPAN></div></DIV>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><I><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: #0000bf; FONT-SIZE: 10pt">Dr Sunil K.Arora<BR>Professor <BR>In Charge- HIV Diagnostic and Disease Monitoring Laboratory &<BR>NACO State Reference Laboratory,</SPAN></I><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: black; FONT-SIZE: 10pt"></SPAN></div></DIV>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><I><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: #0000bf; FONT-SIZE: 10pt">Councillor- Federation of Immunological Societies of Asia-oceania (FIMSA)</SPAN></I><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: black; FONT-SIZE: 10pt"></SPAN></div></DIV>
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<div class=yiv1947419759MsoNormal><I><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: #0000bf; FONT-SIZE: 10pt">Department of Immunopathology<BR>PGIMER, Chandigarh-160 012<BR>Ph.: 0091-172-2755192 (Off)<BR>FAX: 0091-172-2744401, 2745078<BR>Ph:+91-172-4666087 (Res); 9872866087 (cell)<BR>email: <A href="mailto:skarora_in@yahoo.com" rel=nofollow target=_blank ymailto="mailto:skarora_in@yahoo.com">skarora_in@yahoo.com</A> ; <A href="mailto:skarorain@gmail.com" rel=nofollow target=_blank ymailto="mailto:skarorain@gmail.com">skarorain@gmail.com</A> </SPAN></I><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: #0000bf; FONT-SIZE: 10pt"></SPAN></div></DIV>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><B><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: black; FONT-SIZE: 10pt">From:</SPAN></B><SPAN style="FONT-FAMILY: 'Arial', 'sans-serif'; COLOR: black; FONT-SIZE: 10pt"> Clos Joachim <clos@bni-hamburg.de><BR><B>To:</B> Leish-L email post <Leish-l@lineu.icb.usp.br> <BR><B>Sent:</B> Monday, 11 March 2013 8:16 PM<BR><B>Subject:</B> [Leish-l] Fwd: hot spots</SPAN><SPAN style="COLOR: black"></SPAN></div></DIV>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Baghdad', 'serif'; COLOR: #010101; FONT-SIZE: 10pt">Are axenic amastigotes a valid model (and for what?), the first developmental step towards real amastigotes, or just stressed-out promastigotes? Many of us use axenics and swear by them, others are skeptical, others will accept what's inside a macrophage, still others only want amastigotes isolated from something with a fur. I believe we should come to some sort of common language and interpretation. I am somewhere in the middle myself, preferring to do things in macrophages whenever possible.</SPAN><SPAN style="COLOR: black"></SPAN></div>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Baghdad', 'serif'; COLOR: #010101; FONT-SIZE: 10pt">Cheers!</SPAN><SPAN style="COLOR: black"></SPAN></div></DIV></DIV></BLOCKQUOTE></DIV>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><SPAN style="COLOR: black">Am 07.02.2013 um 20:38 schrieb Carlos Costa:</SPAN></div></DIV>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: black">What are the most polemic issues in leishmaniasis?<BR clear=all></SPAN><SPAN style="COLOR: black"></SPAN></div>
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<div style="BACKGROUND: white" class=yiv1947419759MsoNormal><SPAN style="FONT-FAMILY: 'Verdana', 'sans-serif'; COLOR: black">Carlos H. N. Costa, MD, DSc.</SPAN><SPAN style="COLOR: black"></SPAN></div>
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