This is a list of experiments that we would like to do:
| What | Why | Parts |
|---|---|---|
| Point mutate Syk to keep it from being phosphorylated | No phosphorylation = no downstream effects | Syk plasmid |
| Point mutate Lyn to make it autophosphorylate | Control: Even if the BCR complex does not work (eg doesn't detect amyloid), we should still see Syk recruitment | Lyn plasmid |
Check IgM localization using cytometry (see Hombach paper) I believe we use a stained antibody that attaches to igM and see if it sticks to the outside of the cell | IgMkappa IgM heavy CD79a CD79b | |
| Check CD79a/b localization | fluorescently tagged CD79a/b | |
| Check Lyn localization | Lyn-GFP | |
| Check Syk localization | Syk-GFP | |
| Check TEV cleavage | Syk+TEV beta-amyloid CD79-TEVsite-GFP |
does antibody get to membrane?
does cd79A/B get to membrane?
does lyn attach?
does syk get recruited by lyn?
does TEV cleave cut site?
does TEV linked to syk cleave cut site?
-how much does it cut as a function of amount of Syk?
-where should we put molecule to be cleaved?
does beta amyloid trigger release of signal
-get an input-output curve for beta amyloid