Dear Craig,

I agree with the previous posts, but want to add that what the safety cabinet does is to protect your sample from you by shielding it with a laminar air flow. If the inside of the cabinet is not cleaned, the air flow itself will not make any difference. So it is important to wipe surfaces with plenty of ethanol before and after work, sometimes also during work if lengthy procedures. Anyhow, I do not consider the surface itself sterile (not after you move your coat-clad arms over it the first time). If I accidentelly touch the pipette to it I discard it immediately.

Hope these answers helped.

Best,

K

As I understand it, the HEPA filtered air blows out the hood to minimize the chances of airborne bugs getting into the hood.

A regular open TC hood will never be completely sterile and blowing air all day long really doesn't do anything except waste energy and cut down on the filter lifetime.

Dear Craig,

I have been working in category II type safety cabinets for few years now and my answer to your questions reflect my personal experience and opinion.

A cell/tissue culture room will never be 100% sterile nor will the Safety cabinets. The air flow should not take the whole day to stabilize (or at least hours) and if it is doing so there is a problem with the extraction fan. In part if you are experiencing problem with contamination of your cell culture products it could be due to inadequate extraction. In my view leaving culture hood open all day long is not a good general practice and might be against health and safety too!

The outcome of your cell culture (if meaning contamination) may be affected if the entire culture room is not airtight and air filtered.

So it is better to get a working cabinet and put into practice a routine of not leaving cabinets open all day long.

Hope it helps.

H

It typically takes 10-15 minutes for sterile environment to be established after you turned blower on. If you want to avoid waiting 10-15 minutes prior to using the hood every time you need to, it is a good idea to turn the hood on in the morning and keep it running until end of the day. Among other important factors for maintaining sterility are not lifting sash higher than safe operating position usually marked on the sides of the hood and keeping air intake grills in front and behind working surface unobstructed.

Dear Craig,

I agree with the previous posts, but want to add that what the safety cabinet does is to protect your sample from you by shielding it with a laminar air flow. If the inside of the cabinet is not cleaned, the air flow itself will not make any difference. So it is important to wipe surfaces with plenty of ethanol before and after work, sometimes also during work if lengthy procedures. Anyhow, I do not consider the surface itself sterile (not after you move your coat-clad arms over it the first time). If I accidentelly touch the pipette to it I discard it immediately.

Hope these answers helped.

Best,

K

I am agree with Vasily. Certainly, a hood needs at least a yearly certification, when the airflow and the performance of the HEPA filter is checked. Otherwise, run the uv light and airflow for 15 mins, then turn off the uv and you can start working, and the environment will be reasonably sterile.

Indeed, maintaining a constant working flow all the day is required only if you work permanently in the hood. The hood must be eqiped with a sliding door. After use, simpply slide down thsi door, and let a minimal flow in the hood; never completely stop the flow. To check sterility, let a bacteriological dish in the hood for some houres, and then give it to the analytical lab of your local hospital. The sterility, and the quality of the filter can also be checked with a particle counter. Cell culture hood suppliers can do that.

Sterility of athmosphere inside the hood critically depends upon the air moves within the lab. Thus the lab must be somewhat air-tight, and door opening maintained at a minimum. Introducing materials and objects, even packed in sterile enclosure in the working area, is a risk of contamination because coming from a non-sterile zone . The trick is to let these objects (bootles, pipettes, dishes) inside the hood, but the presence of these packages in the working area distubs laminar ait flow. Hard compromise.... A sterilizing heat spot in the working area (gas flame, electrical heating coil) is indirectly useful, when working: this maintain a thermal gradient between the inside and the outside, which contribute to reduce the entry of particles in the working space.

I would also agree with Vasily. In addition I would also allow 10-15 minutes after use to filter the air in the hood of any micro-organisms that the operator has added accidentally, for the benefit of anyone else using the hood. I would also make sure that the surfaces inside the hood are sanitised with 70% ethanol or another suitable substance when turning the hood on and before turning it off (I only mention this as I've known some people not to). I would only leave the hood running all day if I or someone else would be using it throughout the day, though the only thing it wastes is electricity and possibly the filter. It won't improve sterility.

I agree with much of the above and would add check the manual for the intended operation of your hood. Modern hoods have improved in energy efficiency, automated control of the blower etc. Start with the recommendation for your specific hood and proceed from there with what makes sense.

Many usefull comments have already been made... I would like to add just one advise, that it might be helpful to maintain a sterile atmosphere inside the hood by using UV light at the end of the working day (many hoods are equipped with an integrated UV-lamp).

In the lab I am currently working in, we have a scedule for the hoods were everyone can write his/her name in. If during the day there is a gap, we switch the hood to standby mode - usually the last person who has been at the hood switches on the UV for 47min. If after that somebody comes, who didn't write him-/herself to the scedule, he/she switches on the hood again (waiting for 10min) and uses the UV after work again.

OK for UV treatment, but two advices: 1) change regularly the UV lamp, which loses efficiency with time and 2) the hood AND the incubators must be light-tight because UV rays are armful also for the cells you cultivate!

Dear Craig,

It is extremely unusual that one would need to leave the hood switched on all day long to attain a reasonable airflow. This is a waste of electricity and also it will generate a lot of heat in the lab making it unbearable to work (I am not sure how good your cooling system is!), not to mention the constant noise!!

Depending on the model being used and the manufacturer's recommendations, you would need about 5-10 minutes to allow for the air-flow to stabilize.

One of the reasons that your flow cabinet is taking so long to stabilize might be due to blockage in the filter grill (sometimes placed at the back of the hood – see this link: (http://www.invitrogen.com/site/us/en/home/References/gibco-cell-culture-basics/cell-culture-equipment/laminar-flow-hood.html). This grill filter is there to stop any solid materials such as tissues, pipette tips to be sucked up into the HEPA filter system. So please do check it and if it looks dirty then replace it with a clean one and you will find that the hood will only take a short while to stabilize. Be sure to clean and sterilize the hood after you change the filter.

Also, if your hood has perforated surface, then you want to make sure that the surface is not blocked with clutter of bottles, equipment, stands etc., I know many people are extremely lazy to remove things out of the cabinet in case they get “contaminated”. My rule of thumb is that if it is not being used, then it should not be there, and even if it is used, if it obstructs the functioning of the unit, then it should be removed by the user once they are done.

Have a look at the following video link for aseptic techniques: (http://www.sigmaaldrich.com/life-science/cell-culture/learning-center/cell-culture-videos.html).

Thanks, everyone, for your comments. You've confirmed that our cabinet doesn't need to run all day, and that 10 min before use should be sufficient. Now to convince my colleagues...

Our cabinet is professionally cleaned, filters changed and re-certified annually. 70% ethanol, UV treatment, etc. are all things that I do already. Thanks, again, everyone, for confirming what I thought should be standard practice.

Hi Craig!

The only thing I have to add to all comments. I agree with Basil Annaz, don't fill your cabinet with unnecessary material because it tends to block the air-flow depending on the type of the chamber and if there's many different people using the chamber make a schedule for a routine cleaning of them. In my lab there's at least 25 different people using the chambers, so we have a weekly cleaning to clean all the chamber with 70% ethanol in a more consistent manner and check and switch the filters in needed. Also remove the bottom and clean under it because sometimes small paper residues may escape to there and may clogh the filters.

Hope this helps :)

I prefer to keep on the air hood if someone using in the morning than till the remaining day hours. Actually it is a good idea to remain on the air flow especially if lab is busy and prevent from unwanted cleaning and using harmful UV ray every time especially in Asian condition where air contains a lot more dust than other parts of the world. We are keeping on normally. I think this will help you to decide what is good for your lab.

...since almost everything is said, I like to add just two short comments: I didn't like the "10 min" rule, cause its not very precise, so just have a look at your displays in your hood, normally there would be a light or note indicating if the air-flow has reached its ideal value. This might be 10 min but could be less or more.

Second note: don't only clean the surface of the hood, but also underneath the inner aluminum oder steel sheets, you will be surprised what is growing and rotting there. And neither the ethanol nor the UV from above will reach this area. We have a plan to clean this part once a week.

You may find the attached article helpful

Hi Craig

One basic question. Once the LF is switched on in the morning, how it is used by all in the lab? After using the system by a person, it is advisable to wipe and apply UV before the next person start his work. Again if the same person want to handle different type of cells, the procedure is same to avoid cross contamination of cells. It is not clear how this is practiced in your lab. If more than one person use the same hood without cleaning and UV application, it might affect the cell culture outcomes.

Any hood, not only pc2 (biosafety level 2) hoods, is expected to be free from contamination (sterile).

Anil

It is better to wipe the surface with ethanol for each and every time before starting to work in the hood. What ever the cells may be it will be good to clean the surface of the hood right!