Under the Microscope – Tim Kalafut

In his presentation at ISHI 33 this year, Tim Kalafut will describe an interlaboratory DNA interpretation experiment using data from eight forensic biology laboratories and the probabilistic genotyping system STRmix™. Each laboratory contributed twenty mixtures of two to four contributors from their validation studies along with their STRmix™ parameters including STRmix™ kit files and stutters. The extent to which STRmix™ parameters which have been optimized by individual laboratories can be applied to mixture data created in other laboratories was investigated. Previous research concentrated solely on sensitivity studies involving a single type of PCR kit and a small number of laboratories. In this study, we looked at laboratories that used different PCR kits, different numbers of PCR cycles, and different versions of STRmix™. All eight laboratories’ STRmix™ kits were updated to work on STRmix™ version 2.9.0 and all mixtures were evaluated with this version. This interlaboratory study using known sets of donors is a study of the typed called for by the NIST foundation review of DNA Mixture Interpretation.


We chatted with Tim to discuss the impetus for the study, any surprises that arose, and why he’d like to have dinner with Al Capone.



Can you briefly discuss the inter-laboratory probabilistic genotyping study you’ll be presenting at ISHI? What was the impetus for the study and what did it entail?

The idea for this study came about prior to the NIST foundation review draft report in summer of 2021, making this a very timely project. We just wanted to see how STRmix™ performed using different parameters on different data. So we tested the ability of STRmix™ to use validated parameters (STRmix™ “kits”) from eight different STRmix™ labs on one another’s mixtures. In essence, is STRmix™ reliable enough to get concordant results using non-cognate STRmix™ settings/kits/stutters compared to the cognate STRmix™ kits. The similar results from different STRmix™ kits on all those different labs’ mixtures might be useful in addressing some of the NIST draft report concerns related to reliability of PG systems.


How many laboratories participated in the study? What types of samples were analyzed and how were they analyzed?

A total of 8 different labs participated in this study. We asked for 20 mixtures of 2-4 contributors and a range of templates and mixture ratios from each lab. We put no other parameters on our request, as we don’t get to pick and choose our evidence mixtures in forensic DNA casework. We used the input files provided by the labs to match the allele and stutter peaks they analyzed in their own mixtures. The final data table for the portion of the project we will share at ISHI involved over 6000 STRmix™ runs, and each run was compared to a database of the true donors and an additional 10,000 non-donors. This means our study looked at over 61 million comparisons. The actual number is higher due to the number of contributors to each mixture.


Were there any surprises from this study?

The biggest surprise, although surprise is a bit of a strong word, is that we appear to have concordant results for different PCR cycles (28 and 29) and different kits (GlobalFiler and Investigator 24plex. Meaning STRmix™ set up for 28 cycle GlobalFiler data seemed to perform just fine on 29 cycle Investigator 24plex mixtures, and vice versa. Also, any differences between STRmix™ versions seem to be very minimal, although that part of the project hasn’t been fully explored yet.


What tips would you give to someone who is just starting out in the field of forensics, or what is the best advice that you’ve received?

I would advise students that are interested in forensics to get comfortable speaking in front of people. Join a campus club and run for leadership so that you have to speak in front of people. If you play an instrument or sing, find a place to perform. Do whatever it takes to become comfortable in front of an audience. Because you have an audience at court, and at any one moment during the trial, it might feel like half of your audience hates you. Yet you have to be neutral, calm, and professional at all times. If you have no other option, try the Toastmasters International organization.


If you had to pick one thing, what do you enjoy most about your job?

In my current job as an academic, I love the entrepreneurial component of it. I can do any project I want – as long as I find money to pay for it or collaborators to work with. I wasn’t expecting that after 23 years in the crime lab where I had to do either 1) get permission for every special project, or 2) do everything according to protocol. Although you didn’t ask, I had no idea that a new faculty member would be putting in 7-day work weeks of 14-hour days. That’s not exactly a dis-like, but it was unexpected.


If you currently work in the lab, or have in the past, what’s the weirdest thing you’ve ever collected DNA from?

An old case involving a blunt force trauma had a new lead years later, resulting in a search of the woods and a river. A pickaxe was collected from the river in a big old PVC section of pipe, capped at both ends. (The handle was sawed off so both the handle and the head would fit into the pipe.) It was really, really heavy because it was full of water. Turns out, because the axe was in the water, they wanted to capture any DNA that might have been washed off. Needless to say, I didn’t collect any DNA from it.


If you could have dinner with anyone (dead or alive), who would it be? Why?

Al Capone. To find out what was really in that vault….


If you’ve attended ISHI in the past, what do you most enjoy about coming to the conference? If you haven’t, what are you most looking forward to?

Catching up with friends from around the world.


What’s one thing that others may not know about you?

I got shot in the head with a bow and arrow as a kid.