Re: [ccp4bb] insect cell media
Hi, We are using a modified version of ISFM with 10 g/L glucose, based on a paper by Maiorella et al. from around 1988 or 1989. This is a medium that is based on IPL-41, which we purchase as a powder, and supplemented with ultrafiltered yeastolate and lipids. It is completely protein-free. It is exceptionally cheap but does require a little more work than commercial media to prepare (you have to emulsify the lipids yourself). The medium costs us $3.92 per L before filter sterilization, and $5.26 per L after. We use 2 Stericup 500mL PES 0.2μm (Express Plus) bottle-top filters for each 10 L prep of liquid media. Note that these prices include institutional discounts for some components that may or may not apply to you. I can send you our protocols if you are interested. -Tom --- Thomas Cleveland Leahy Lab Biophysics Biophysical Chemistry Department Johns Hopkins University School of Medicine On Mon, Jun 7, 2010 at 7:18 AM, Chris Ulens chris.ul...@med.kuleuven.be wrote: Hi, I would like to get some feedback on the type of media that people are using for insect cell culturing. We have happily made a transition last year from using Invitrogen's SFM medium and cellfectin to Insect-Xpress (Lonza) and polyethyleneimine for transfection. We are moving several protein targets to large-scale cultures and would consider cost-cutting alternatives. For example, Invitrogen and Thermo Sci both offer media in powder form at an attractive price. Has anyone made a systematic comparison of these media? Any particular recommendations regarding powder media? Thanks. -Chris --- Chris Ulens, Ph.D. Lab of Structural Neurobiology Department of Molecular Cell Biology Campus Gasthuisberg, ON1 Herestraat 49, PB 601 B-3000 Leuven Belgium e chris.ul...@med.kuleuven.be t +32 16 345812 f +32 16 345699 w http://www.xtal.be
Re: [ccp4bb] insect cell media
We have happily made a transition last year from using Invitrogen's SFM medium and cellfectin to Insect-Xpress (Lonza) and polyethyleneimine for transfection. We are moving several protein targets to large-scale cultures and would consider cost-cutting alternatives. For example, Invitrogen and Thermo Sci both offer media in powder form at an attractive price. Has anyone made a systematic comparison of these media? Any particular recommendations regarding powder media? Last I checked, no manufacturer offered serum-free powedered medium. Fetal calf serum is very expensive - once you count in its cost, the overall price of the media becomes comparable. If your protein is secreted, serum-containing media are not even a realistic choice. If it is intracellular, buying dry medium and adding FBS work well: is a lot more hassle, is about 20% cheaper, and usually gives slightly higher titers and expression levels. Of all the common media we compared, TNM-FH works best for us with Sf9 cells. Switching to dry medium also means some initial investment: bottles, filtration equipment and filters. So it all pays off only of you need large scale cultures long-term. If you are aware of a dry serum-free insect cells medium, please let me know - I'd love to try it. Dima
Re: [ccp4bb] insect cell media
Hi, well, actually this is availalbe as powder (HYClone), as it says on the page.. (we have it made in our media kitchen on a regular basis, but not for huge scale... so i dont know about the prices..) https://www.thermoscientific.com/wps/portal/ts/products/detail?navigationId=LA11074__10347categoryId=82051productId=11960716 HTH, Tommi On Jun 7, 2010, at 5:52 PM, Dima Klenchin wrote: We have happily made a transition last year from using Invitrogen's SFM medium and cellfectin to Insect-Xpress (Lonza) and polyethyleneimine for transfection. We are moving several protein targets to large-scale cultures and would consider cost-cutting alternatives. For example, Invitrogen and Thermo Sci both offer media in powder form at an attractive price. Has anyone made a systematic comparison of these media? Any particular recommendations regarding powder media? Last I checked, no manufacturer offered serum-free powedered medium. Fetal calf serum is very expensive - once you count in its cost, the overall price of the media becomes comparable. If your protein is secreted, serum-containing media are not even a realistic choice. If it is intracellular, buying dry medium and adding FBS work well: is a lot more hassle, is about 20% cheaper, and usually gives slightly higher titers and expression levels. Of all the common media we compared, TNM-FH works best for us with Sf9 cells. Switching to dry medium also means some initial investment: bottles, filtration equipment and filters. So it all pays off only of you need large scale cultures long-term. If you are aware of a dry serum-free insect cells medium, please let me know - I'd love to try it. Dima Tommi Kajander, Ph.D. Structural Biology and Biophysics Institute of Biotechnology University of Helsinki Viikinkaari 1 (P.O. Box 65) 00014 Helsinki Finland p. +358-9-19158903 tommi.kajan...@helsinki.fi
Re: [ccp4bb] insect cell media
I mentioned to to Chris already, but we use nothing but HyClone SFX-Insect powder. We make 20-30 L batches, sterilize with a large peristaltic pump and a disposable Millipak filter from Millipore. We never have contamination problems that are due to preparing our own media from powder. We buy 10L bottles of powder. This media works great for baculovirus, stable cells, Sf9, Sf21, Tn5. We prefer it over Invitrogen (why have separate media for Sf9 and Tn5 cells?), and it's cheaper too! We no longer use serum media for anything (virus production and amplification, all goes fine in serum free. Serum helps storing viral stocks for long periods). It's ~$26 per L, plus $1-$2 in filter costs (a $30 filter will sterilize ~30 L of media). So it comes to about ~$28/L, I haven't priced out alternative products recently, but I think liquid from Invitrogen runs around $50/L? We also use the peristaltic pump for tangential flow filtration, it's pretty heavy duty (tubing is like 5/8 thick). I think that help make the filtering efficient. Bottle top filters won't work because they foul to quickly, same is try of 4.5 cm filter discs that you can insert into an in-line filter, they clog after 1 L or so. You need the 'asymmetric' filter material offered by Millipore in the Millipak or Steripak filter (the later is the filter you see attached to a MilliQ water dispenser, it can be autoclaved 3X they claim) Hope that helps, Nat On Mon, Jun 7, 2010 at 10:59 AM, Tommi Kajander tommi.kajan...@helsinki.fi wrote: Hi, well, actually this is availalbe as powder (HYClone), as it says on the page.. (we have it made in our media kitchen on a regular basis, but not for huge scale... so i dont know about the prices..) https://www.thermoscientific.com/wps/portal/ts/products/detail?navigationId=LA11074__10347categoryId=82051productId=11960716 HTH, Tommi On Jun 7, 2010, at 5:52 PM, Dima Klenchin wrote: We have happily made a transition last year from using Invitrogen's SFM medium and cellfectin to Insect-Xpress (Lonza) and polyethyleneimine for transfection. We are moving several protein targets to large-scale cultures and would consider cost-cutting alternatives. For example, Invitrogen and Thermo Sci both offer media in powder form at an attractive price. Has anyone made a systematic comparison of these media? Any particular recommendations regarding powder media? Last I checked, no manufacturer offered serum-free powedered medium. Fetal calf serum is very expensive - once you count in its cost, the overall price of the media becomes comparable. If your protein is secreted, serum-containing media are not even a realistic choice. If it is intracellular, buying dry medium and adding FBS work well: is a lot more hassle, is about 20% cheaper, and usually gives slightly higher titers and expression levels. Of all the common media we compared, TNM-FH works best for us with Sf9 cells. Switching to dry medium also means some initial investment: bottles, filtration equipment and filters. So it all pays off only of you need large scale cultures long-term. If you are aware of a dry serum-free insect cells medium, please let me know - I'd love to try it. Dima Tommi Kajander, Ph.D. Structural Biology and Biophysics Institute of Biotechnology University of Helsinki Viikinkaari 1 (P.O. Box 65) 00014 Helsinki Finland p. +358-9-19158903 tommi.kajan...@helsinki.fi
Re: [ccp4bb] insect cell media
For large amount of media, it's actually cheaper to buy liquid media in bags when factor in the cost of labor and water. MilliQ water may not be low endotoxin. --Chun -Original Message- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Nathaniel Clark Sent: Monday, June 07, 2010 11:00 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] insect cell media I mentioned to to Chris already, but we use nothing but HyClone SFX-Insect powder. We make 20-30 L batches, sterilize with a large peristaltic pump and a disposable Millipak filter from Millipore. We never have contamination problems that are due to preparing our own media from powder. We buy 10L bottles of powder. This media works great for baculovirus, stable cells, Sf9, Sf21, Tn5. We prefer it over Invitrogen (why have separate media for Sf9 and Tn5 cells?), and it's cheaper too! We no longer use serum media for anything (virus production and amplification, all goes fine in serum free. Serum helps storing viral stocks for long periods). It's ~$26 per L, plus $1-$2 in filter costs (a $30 filter will sterilize ~30 L of media). So it comes to about ~$28/L, I haven't priced out alternative products recently, but I think liquid from Invitrogen runs around $50/L? We also use the peristaltic pump for tangential flow filtration, it's pretty heavy duty (tubing is like 5/8 thick). I think that help make the filtering efficient. Bottle top filters won't work because they foul to quickly, same is try of 4.5 cm filter discs that you can insert into an in-line filter, they clog after 1 L or so. You need the 'asymmetric' filter material offered by Millipore in the Millipak or Steripak filter (the later is the filter you see attached to a MilliQ water dispenser, it can be autoclaved 3X they claim) Hope that helps, Nat On Mon, Jun 7, 2010 at 10:59 AM, Tommi Kajander tommi.kajan...@helsinki.fi wrote: Hi, well, actually this is availalbe as powder (HYClone), as it says on the page.. (we have it made in our media kitchen on a regular basis, but not for huge scale... so i dont know about the prices..) https://www.thermoscientific.com/wps/portal/ts/products/detail?navigationId= LA11074__10347categoryId=82051productId=11960716 HTH, Tommi On Jun 7, 2010, at 5:52 PM, Dima Klenchin wrote: We have happily made a transition last year from using Invitrogen's SFM medium and cellfectin to Insect-Xpress (Lonza) and polyethyleneimine for transfection. We are moving several protein targets to large-scale cultures and would consider cost-cutting alternatives. For example, Invitrogen and Thermo Sci both offer media in powder form at an attractive price. Has anyone made a systematic comparison of these media? Any particular recommendations regarding powder media? Last I checked, no manufacturer offered serum-free powedered medium. Fetal calf serum is very expensive - once you count in its cost, the overall price of the media becomes comparable. If your protein is secreted, serum-containing media are not even a realistic choice. If it is intracellular, buying dry medium and adding FBS work well: is a lot more hassle, is about 20% cheaper, and usually gives slightly higher titers and expression levels. Of all the common media we compared, TNM-FH works best for us with Sf9 cells. Switching to dry medium also means some initial investment: bottles, filtration equipment and filters. So it all pays off only of you need large scale cultures long-term. If you are aware of a dry serum-free insect cells medium, please let me know - I'd love to try it. Dima Tommi Kajander, Ph.D. Structural Biology and Biophysics Institute of Biotechnology University of Helsinki Viikinkaari 1 (P.O. Box 65) 00014 Helsinki Finland p. +358-9-19158903 tommi.kajan...@helsinki.fi
