***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi again I asked Thermo Fisher and got this: -------------------------------------- Phenol red (and other cyclic compounds) has been shown to increase background fluorescence (its peak is 440 nm), which can create issues for users depending on the channels being used. One such reference is https://link.springer.com/protocol/10.1385/1-59259-826-9:395. Hence if using colors in that spectrum, it is best to use phenol red free medium. Additionally phenol red's cyclic nature is similar to that of estrogen, so it can bind to and activate ERs of estrogen sensitive cells (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC323325/). If doing work in those areas it is also a good idea to remove phenol red. John Fisk, Ph.D. Technical Applications Scientist Life Sciences Solutions Group Thermo Fisher Scientific T (NA) 800-955-6288 | (EMEA) 00-800-5345-5345 [log in to unmask] | thermofisher.com ------------------------------------------------- The ref given by John from Thermo Fisher shows a 20% increase in background fluorescence in the green and red channel when using Phenol red compared to medium without. There is also an interesting information about Estrogen which I did not know. Absolutely nothing about quenching fluorescence. However, I saw that the measurements were done on plastic dishes. I did measure background fluorescence in the green and red channels with and without Phenol red some years ago and never found any difference. I wonder if one would get the same results on glass bottom dishes. Pawley's confocal handbook (p 361) mentions that the problem with phenol red is fluorescence quenching but there is no ref. I would say that the question is not fully solved... Med vänlig hälsning / Best regards Sylvie @@@@@@@@@@@@@@@@@@@@@@@@ Sylvie Le Guyader, PhD Live Cell Imaging Facility Manager Karolinska Institutet- Bionut Dpt Hälsovägen 7C, 14157 Huddinge, Sweden mobile: +46 (0) 73 733 5008 LCI website Follow our microscopy blog! -----Original Message----- From: Confocal Microscopy List <[log in to unmask]> On Behalf Of Model, Michael Sent: 15 September 2021 18:41 To: [log in to unmask] Subject: Re: EXT: Phenol red in imaging medium ***** To join, leave or search the confocal microscopy listserv, go to: https://eur01.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.umn.edu%2Fcgi-bin%2Fwa%3FA0%3Dconfocalmicroscopy&data=04%7C01%7Csylvie.le.guyader%40KI.SE%7C6e92ce91403941f654bc08d978684199%7Cbff7eef1cf4b4f32be3da1dda043c05d%7C0%7C0%7C637673211709136758%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C3000&sdata=AJdzBTx9NAdq58%2FJHaNmigY7zDnlczMXZY25yE18MsA%3D&reserved=0 Post images on https://eur01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.imgur.com%2F&data=04%7C01%7Csylvie.le.guyader%40KI.SE%7C6e92ce91403941f654bc08d978684199%7Cbff7eef1cf4b4f32be3da1dda043c05d%7C0%7C0%7C637673211709146746%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C3000&sdata=C20tpeJFJoe2rIPZSOj7ECu4kU9QJPjbGjRXtSDWUGo%3D&reserved=0 and include the link in your posting. ***** It seems to have a strong absorbance at 550 only at alkaline pH (https://eur01.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.researchgate.net%2Fpublication%2F221925346_Plastic_Optical_Fiber_pH_Sensor_Using_a_Sol-Gel_Sensing_Matrix%2Ffigures%3Flo%3D1&data=04%7C01%7Csylvie.le.guyader%40KI.SE%7C6e92ce91403941f654bc08d978684199%7Cbff7eef1cf4b4f32be3da1dda043c05d%7C0%7C0%7C637673211709146746%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C3000&sdata=DdJTtTEN163wfYxI5naW9kXsatMUWu56nufjAwlsajw%3D&reserved=0). But if left in the room, bicarbonate-based media will turn alkaline, so it is possible that some quenching by energy transfer occurs. Mike -----Original Message----- From: Confocal Microscopy List <[log in to unmask]> On Behalf Of Sylvie Le Guyader Sent: Wednesday, September 15, 2021 10:51 AM To: [log in to unmask] Subject: EXT: Phenol red in imaging medium ***** To join, leave or search the confocal microscopy listserv, go to: https://eur01.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.umn.edu%2Fcgi-bin%2Fwa%3FA0%3Dconfocalmicroscopy&data=04%7C01%7Csylvie.le.guyader%40KI.SE%7C6e92ce91403941f654bc08d978684199%7Cbff7eef1cf4b4f32be3da1dda043c05d%7C0%7C0%7C637673211709146746%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C3000&sdata=5M%2Bjk%2BkOqD32MwJE4Dw%2Ffv3N5ZGKqkZ3kbO69frI7rc%3D&reserved=0 Post images on https://eur01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.imgur.com%2F&data=04%7C01%7Csylvie.le.guyader%40KI.SE%7C6e92ce91403941f654bc08d978684199%7Cbff7eef1cf4b4f32be3da1dda043c05d%7C0%7C0%7C637673211709146746%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C3000&sdata=C20tpeJFJoe2rIPZSOj7ECu4kU9QJPjbGjRXtSDWUGo%3D&reserved=0 and include the link in your posting. ***** Dear all I have heard that the reason why Phenol red is avoided in cell culture media used for imaging is that it quenches fluorescence in some (at least the green) spectrum. I cannot find any reference about this. 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