What "assays" are, some ways they're classified, & some key things about them (biochemistry focus)

“Assay” is just a fancy word for an experiment where we measure something. And by “something” we’re talking in really broad terms - that something can be a lot of different things. In the simplest sense, we’re measuring numbers of a specific molecule (concentration assays). But we can also, especially in biochemistry, measure amount of “do-ability”. How much ability is there to bind some other molecule (binding assay)? Or how much ability is there for some enzyme to do whatever that enzyme does (enzyme activity assay)?
blog: bit.ly/biochem...
Those different “something’s” require different types of assays. And even within the same type of something, which assay you use will depend on what molecules you’re working with and what equipment you have available. There is typically not just one assay you can use. Instead, there are lots of assays that can provide the same, or at least similar, information - although each will have strengths and weaknesses. So I encourage you to learn as much as you can about different assays that are commonly used. Not only will this help you design and carry out your own experiments, but it will also help you evaluate the results of assays others have done (as reported in papers, etc.).
In an assay you often measure the “something” indirectly, such as through a dye, fluorescence, etc. And you often systematically vary concentrations in order to get more information (binding constants, activity rates, etc.).
Common assays you might come across in biochemistry include:
- Various “protein assays” to measure protein concentration - Bradford assay, BCA assay, Lowry assay etc.
- Various binding assays to measure binding strength - SPR (surface plasmon resonance), ITC (isothermal calorimetry), ELISA (enzyme-linked immunosorbent assay), EMSA (electrophoretic mobility shift assay), slot-blots, etc.
- Various enzyme assays to measure enzyme activity - lots of different ones because enzymes do lots of different things so for each enzyme you need a different type of assay!
In addition to classifying assays based on what they’re measuring, or what equipment they use, we can classify them based on what type of data they give us: quantitative, qualitative, or semi-quantitative.
When you see “quantitative,” think “numbers.” Or I guess in this case, when you see numbers think quantitative! In terms of what an assay can tell you, you can basically think of things as:
- Qualitative: is the thing present? (Yes/no)
- E.g a pH indicator can tell you whether a solution is acidic (if there are a there a lot of protons (definition of acidity) the solution will change color)
- Semi-qualitative: roughly how much of the thing is present? (Ballpark figures, high/med/low, etc.)
- E.g. pH test strips can estimate how acidic the solution is
- Quantitative: exactly how much of the thing is present? (At least within the limits of the assay)
- E.g. a pH probe can tell you precisely what the pH is
more on protein concentration assays: bit.ly/proteinm...
more on binding assays: bit.ly/bindinga...
more on enzyme activity: bit.ly/enzymeac... & • Specific activity & en...
more details on various assays (and other biochemistry techniques): bit.ly/labtechn...
more terminology: bit.ly/bumbling... & www.youtube.co...
 
more about all sorts of things: #365DaysOfScience All (with topics listed) 👉 bit.ly/2OllAB0 or search blog: thebumblingbioc...

[@Opinionated05]

Thank you so much!

[@kenpalmer3315]

Thank you - you simplify complex concepts!

[@thebumblingbiochemist]

So glad I could help!

[@jahnavee_palsodkar]

this is actually so helpful, thank you!!

[@thebumblingbiochemist]

Awesome!

[@user-uf1ke8md4k]

Hello Dr. Bibel, I just wanted to say thank you, as your videos have really helped me with my research as an undergraduate student.
Are you able to make a video on scintillation proximity assays or peptide spot assays?
Update: your "radiometric kinase assay with scintillation counting" helped with understanding the scintillation proximity assay. Any information on a peptide spot assay would be much appreciated! Thank you!

[@thebumblingbiochemist]

Sorry but I don't anticipate doing one of those in the near future. Good luck!

[@gasparomarsala6078]

thank you

[@krishnasai1288]

Please make a video on BCA assay

[@thebumblingbiochemist]

I hope to in the future - for now I do have some content on it here: bit.ly/proteinmeasuring

[@strugglingcollegestudent]

Who else is here studying for their biochem exam

[@thebumblingbiochemist]

best of luck! hope some of my content can help!