MAKING CELL CULTURE MEDIA

I. INTRODUCTION
The purpose of this exercise is to acquaint you with the way in which the media used to grow your cell cultures is prepared. The medium will be prepared by you for your "laboratory". That is the way it is done in most cell culture laboratories and it is valuable experience for you to know how to do this if you should be required to do so in the "real world". The procedure is easy to do, the information for making the medium is always contained either on the packet or in the package containing the dried powdered medium. If you are using already reconstituted medium, it will usually come from the manufacturer as either 1X or as 10X (concentrated) where you would then make the necessary dilution and use the medium.

 II. MATERIALS
Glass distilled or deionized water Powdered media - F-12 (two labs will make this medium), EBME, DME/F-12; Filters for sterilization; Collection bottles, Storage bottles, 1N NaOH, 1N HCl, Bovine serum (either fetal, calf or adult).

 III. PROCEDURE
Each of you will make 1L of the medium you will ultimately use for the cells with which you are going to be working. Follow the procedure on the packet of media given to you or in the instructions contained with the small bottles of powdered media. The media you will be making today will be media you will be using throughout the course in planting and preparing cells from one exercise to the other. You will be responsible for maintaining an inventory of the medium you are using. Thus, be certain that you always have, on hand, sufficient medium for the next and subsequent experiments. You will NOT be permitted to borrow medium from the next lab (borrowing any sort of liquid material from another laboratory always leaves you open to potential contamination as you can never be certain of someone else's aseptic technique)--YOU ARE RESPONSIBLE FOR MAINTAINING THE MATERIALS NEEDED BY YOUR LAB. When you are running low on media, "order" powdered medium from the T/A and make up a fresh stock for your lab.

 IV. DISCUSSION
1. What did you learn in this process? You will also be responsible for setting up the cultures for succeeding experiments and so it is important for you to read ahead for what will be needed. One sure way of success is to ALWAYS have one to two bottles of your cells ready each week in case they are needed. Therefore, each week designate another person to carry out the passaging of the cells. (We realize that this is really a breach of technique but, in this situation, it is unavoidable.  We have to rely on each member of the lab to do his/her best.)
2. Look in the catalogs provided and see how the medium you will be using is provided by the manufacturer. Provide us with the specific catalog number of the medium you prepared and the serum you used.

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