I have Microarray data with 3 sample each in uninfected_48hrs, infected_48hrs, uninfected_60hrs and infected_60hrs.

```
#3 sample infected_48h; 3 sample Uninfected_48h; 3 sample infected_60h & sample Uninfected_60h
#First design matrix looks like this:
infected_48h infected_60h Uninfected_48h Uninfected_60h
Uninfected_48h.1 0 0 1 0
Uninfected_48h.2 0 0 1 0
Uninfected_48h.3 0 0 1 0
infected_48h.1 1 0 0 0
infected_48h.2 1 0 0 0
infected_48h.3 1 0 0 0
uninfected_60h.1 0 0 0 1
uninfected_60h.2 0 0 0 1
uninfected_60h.3 0 0 0 1
infected_60h.1 0 1 0 0
infected_60h.2 0 1 0 0
infected_60h.3 0 1 0 0
attr(,"assign")
[1] 1 1 1 1
attr(,"contrasts")
attr(,"contrasts")$sample.levels
[1] "contr.treatment"
# Second design matrix
Uninfected_48h infected_48h infected_60h Uninfected_60h
Uninfected_48h.1 1 0 0 0
Uninfected_48h.2 1 0 0 0
Uninfected_48h.3 1 0 0 0
infected_48h.1 1 1 0 0
infected_48h.2 1 1 0 0
infected_48h.3 1 1 0 0
uninfected_60h.1 1 0 0 1
uninfected_60h.2 1 0 0 1
uninfected_60h.3 1 0 0 1
infected_60h.1 1 0 1 0
infected_60h.2 1 0 1 0
infected_60h.3 1 0 1 0
attr(,"assign")
[1] 0 1 1 1
attr(,"contrasts")
attr(,"contrasts")$SKN_group_int
[1] "contr.treatment"
# Which design matrix is prefer in this scenario first design matrix or second design matrix?
```

Which design matrix should I used in this case and why? I am new to this design matrix concept. Please do comment and suggest thank you.

Something is not right, here. Please show all prior code, and paste samples of your metadata, if possible. Thank you.

I was curious on which design matrix should I used/based on if I have 12 samples (3 infected_48hr, 3 uninfected, 3 infected_60hr and 3 uninfected_60hr). So I didn't paste the prior code. I think I should used the first design matrix.

Sure, and yes, you can now conduct individual comparisons via

`makeContrasts()`

and`contrasts.fit()`