Experimental Temperature measurement for Model Validation
Cell suspensions are prepared as described before (Dopp, Jo, & Reuel, 2019). In brief, starter culture is prepared by taking 20 mL of LB media (prepared by protocol outlined in [2]) and a cell pick of BL21 DE3 Star. This starter culture is left to grow overnight and used to inoculate 1 L growth media of 2x YTPG (prepared by protocol outlined in [2]) in a 2.5 L Tunair flask. The cells are harvested after 4 hours. The cell pellets are resuspended in S30 buffer, centrifuged (at 3000g for 20 minutes in 4C temperature) and pelleted again. These cells are mixed with equal volume of S30 buffer and vortexed to produce cell suspension which is aliquot in tubes.
The sonication tubes are placed in an ice bath containing ground ice in water, stabilized with a clamp. The sonication tip is dipped in the suspension 0.5 cm from top as per the optimum depth found from this study (Fig 2). A k-type probe data logger thermometer (Fluke TC-2000) is used for temperature measurement and logging. The probe head was carefully held manually inside the sonication tube so that it did not touch the tube wall or the sonication tip due to vibration. The amplitude and pulse settings were specified and the sonication and data logging was started at once. The experiments were conducted with different power setting, pulse setting, tip radius, tubes and initial temperatures to observe the experimental response in different conditions and validate the finite element model. For each of the experimental runs, simulations were also conducted using the model inputs. The experimental runs were -
  1. 1.5 mL microcentrifuge tube with 1.5 mL sample & 50% amplitude 3mm tip (corresponding to 5.5 Watt) and 10 second on and 10 second off pulse, initial temperature 1.6 C. (Fig 3a)
  2. 1.5 mL microcentrifuge tube with 1.5 mL sample & 50% amplitude 3mm tip (corresponding to 5.5 Watt) and 20 second on and 20 second off pulse, initial temperature 3.4 C. (Fig 3b)
  3. 1.5 mL microcentrifuge tube with 1.5 mL sample & 25% amplitude 3mm tip (corresponding to 1.73 Watt) and 10 second on and 10 second off pulse, initial temperature 1.1 C. (Fig 3c)
  4. 5 mL microcentrifuge tube with 5 mL sample & 50% amplitude 6mm tip (corresponding to 12.5 Watt) and 20 second on and 20 second off pulse, initial temperature 14.4 C. (Fig 3d)