20.109(F16):Test comet chip loading variables (Day2)
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Contents
Introduction
experimental design...
Protocols
Part 1: Design experiment to optimize comet chip loading
You will design two experiments that test variables associated with loading cells into the microwells of your comet chip. In the first experiment you will consider the role of cell number. Specifically, how many cells should be added to each well to ensure the majority of the microwells are loaded? In addition, how many cells should be loaded into each microwell? The second experiment will interrogate the length of time needed for loading. In the comet chip that you prepared during the previous laboratory class there is space for you to complete two experiments, each with two conditions.
Experiment 1: Cell number
- How many cells should be loaded into each microwell?
- Consider the amount of DNA that is carried by a single mammalian cell and the detection limit provided by the SYBR gold DNA stain that will be used in your experiments. Also, use the data you collected during the Orientation exercise to determine how many cells can fit into a single microwell based on the dimensions provided above.
- When you know how many cells you want to load into each microwell, move on to the next question.
- How many cells should be added to each well such that the desired number of cells are loaded into the microwells?
- Consider the likelihood that every cell you add to the well will fall into a microwell. Perhaps calculate the surface area of the bottom of a well and compare this to the size of the cells as you consider this question.
- Now that you have an idea as to the number of cells that are ideal for loading, consider how you will test this in your experiment.
Experiment 2: Loading time
Part 2: Load comet chip
Reagents
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