Dear Chhavi,

the only recommendation is to avoid serum with BEAS-2B since they can undergo differentiation which in turn may affect your results. For the coating of the flask I reckon you are already managing. One option is to use cell-bind flasks/petri dishes which work perfectly fine and without the need of coating.

Maurizio

Hi Chhavi,

i don´t know how you wanna use your BEAS-2B cells, but in our lab we cultivate these cells by just seeding them in T75 flasks with DMEM (+P/S and FBS) without coating the flasks before and it always worked. Even in multiwell-plates or smaller flasks.

the same For me. I've always cultured them without coating. But uRPMI medium.

I use DMEM:F12 or HAM's F12 medium both with P/S and FBS to culture BEAS-2B without coating. Moreover the RPMI, HAM, DMEM or other medium are very very less expensive than BEGM Kit.

But if you want to do the coating I think you could dissolve fibronectin, collagen and albumin in BEBM using a volume that cover entirely the flask surface and then leave the medium in the flask overnight. Another tip is to contact the provider of BEGM and ask them the technical guidelines to do the coating

Thanks all for your valuable suggestions.

Miss gupta u can purchase the surface treated flask also, y r u taking pain.

Do I have to add  polyvinylpyrrolidone to trypsin solutions during trypsinization?

http://www.sigmaaldrich.com/catalog/product/sigma/95102433?lang=en®ion=US

This is pretty helpful.. but I feel like you probably have it under control now.

I am starting with BEAS-2B now and I had the same problem. Neither ATCC nor Sigma writes that collagen almost insoluble in medium. I have found this suggestion. Dissolve collagen in 0.1 M acetic acid and sterilize it by incubation with chloroform (filtration is not recomended) (http://www.sigmaaldrich.com/technical-documents/articles/biofiles/collagen-product-protocols.html). So far it seems it works.

I wish to start my work with this cell line. It would be really great if anyone could help with the following questions:

Does culturing the cells in a non coated flask cause any changes in morphology or other characteristics of the cell line?

At what percent is FBS used for routine maintenance?

While freezing, do we need to add PVP as mentioned on ATCC website or routine freezing medium with complete growth medium and DMSO is sufficient?

@Chhavi: Please let me know what worked best for you.

Thank you

I just use DMEM F12 medium. The cell grow well. i used 10% fbs. I just use 10% DMSO for cell storage

I've used the BEGM kit and cultured cells with the coating and without coating. I didn't see any difference in how they grew so I continued using them without coating the flasks. I have also grown the cells in LHC-9 medium and with KGM-Gold from Lonza. The KGM-Gold is the best for me.

Hi, you can find this information on ATCC; https://www.lgcstandards-atcc.org/Products/All/CRL-9609.aspx?geo_country=ch#culturemethod

See Flask coating at middle-down of the page.

Dear Chhavi,

the only recommendation is to avoid serum with BEAS-2B since they can undergo differentiation which in turn may affect your results. For the coating of the flask I reckon you are already managing. One option is to use cell-bind flasks/petri dishes which work perfectly fine and without the need of coating.

Maurizio

Hello

I would be starting my work with the BEAS-2B cell lines. From what ATCC recommends, we should use BEGM but I have read researchers working with the DMEM, RPMI and KGM media for culturing. Does the choice of media effect the growth and response profile/ morphological changes in the growth of cells?

I see a lot of experts here recommending the use of BEAS-2B without pre-coating of flasks/plates. Does it effect cell count and assay results in any way?

Thanks in advance!

Hello

can anyone tell me how to make complete medium with lhc-9 medium??

i want to start my work in beas 2b cell line..can anyone plz help me out/..how to make complete medium?? do we have to add fbs and antibiotic solution or not??

Dear Shahana Parveen

LHC-9 is already complete! Do not add serum (it is a serum free medium) neither antibiotics because they are already there!

We have tried not to pre-coat cultivation surfaces for BEAS-2B cells but it did not work well for us. Now we use coating only with colagen solution and it works quite nice to us.

@ Diego Baderna Thanku for the reply sir.

I thought i would add my experience here if new BEAS-2B users visit.

After using BEAS-2B for several years now i have found:

-BEGM is the best media for these cells in my opinion. Unfortunately it is the most expensive too. No other additives are needed. Be aware it contains antibiotics in the bullet kit if you are looking to do bacterial-based experiments.

-LHC-9 serum free will work OK, but produces a different cell morphology to BEGM. They appear to produce less tight junctions between cells and i would not personally recommend this media. Growth rate and adherence seem reduced. I have not done further testing with LHC-9.

-It is clear now the addition of FBS to any BEAS-2B culture media (RPMI, DMEM etc) produces significantly altered factors and cell phenotype and should always be avoided for scientific integrity. However these changes seem to occur over a week to fortnight of time.

-These cells do not adhere easily to glass. A number of different coatings are available. I would suggest leaving your coating of choice to dry overnight. Seed your BEAS-2B and let them grow overnight. If you continue to have issues, add 10% FBS to the media overnight. FBS contains proteins that help adhesion. But i strongly recommend removing FBS-media 24-48 hours before any exposure as i found it altered the cell response to my exposures.

-Coating is not required for routine cell culturing in cell-culture treated plastic flasks. We use Corning T75 flasks with CellBind. The same for our plates.

-The routine addition of antibiotics to cell culture media is also not recommended, but should not cause any changes to the cell.

Cheers,

can you please explain how to prepare the cryo medium for beas 2 b cells??

according to the atcc its complete medium containing basal medium and single quots supplemented with 1 % polyvinylpyrrolidone (pvp) and 7.5 % DMSO

i have pvp , but its in powder form.. how should i add it to make cryo medium.

--should i add 1 % in medium and mix it.. but here i have i have concern whether it will be soluble in media or not?

--or d i have to dissolve pvp in water and then add accordingly???

also in atcc its given to not to filter complete media whereas in lonza data sheet its written we can filter medium if we want.

@shahana Parveen

I routinely use 10% DMSO, 10% FBS and 80% BEGM to freeze store BEAS-2B.

If you want to use PVP, mix to 1% final volume whatever way you choose to. Mixing into cell storage media seems most logical to me.

Either way ensure you are following a correct freezing protocol using a cell freezing chamber/ container.

I have used both filtered and unfiltered BEGM for routine culture and saw no difference.

I hope this helps,

thanks..