3D printing procedure.
1. Download 3D print files of the Vent-Lock splitters, flow regulators, and manometer adaptors. Print files were generated by CAD drawings (SolidWorks, Dassault Systèms) and converted into G-code using the printer’s accompanying software package (PreForm, Formlabs). 3D Print Files are available upon request.
2. Open 3D print files in Solidworks
3. Print via stereolithography (Form 2, Form 3, or Form 3B, Formlabs) at 50 um layer resolution, using surgical guide resin (Surgical Guide, Formlabs). Generate support structures through PreForm where needed. Orient components in such a way that crucial surfaces such as threads or O-ring ledges are not impacted by support structures.
4. Post-process the prints by washes (2 cycle with 15 min per cycle) in >99.5% isopropyl alcohol (CAS Number: 67-63-0, Sigma Aldrich), followed by air-drying at 22oC for 30 minutes, and post-cure for 30 minutes with heat 60 oC for the Form 2 printer and 70 oC for the Form 3B printer at 405 nm of light (Form Cure, Formlabs).
5. If printing the FloRest, insert O-rings (E1000-212/AS568-212, O-Rings EPDM, FDA EPDM, Marco Rubber & Plastics, Seabrook, New Hampshire, USA) to improve sealing. Production via fused deposition modeling (FDM) (e3d, BigBox3D Ltd, Oxfordshire, UK; Little Monster, Tevo 3D Electronic Technology Co. Ltd, Zhanjiang, China)
6. If printing via FDM, use PETG Filament (PETG 3D Printer Filament, FilaMatrix, Virginia, USA). Set print settings to 0.2mm layer height with 30% infill, nozzle temperature of 250 oC, and bed temperature of 70 oC; generate supports from the build platform, with no interior supports.
1. 3D-printed parts produced from surgical guide resin can be sterilized by dry vacuum autoclave (Sr 24C Adv-PlusTM, Consolidated Sterilizer Systems, Boston, Massachusetts, USA), 3 cycles at 120.0 oC, 20 minutes sterilization time and 20 minutes dry time.
2. 3D-printed parts produced from surgical guide resin can be sterilized by soaking in >99.5% isopropyl alcohol (CAS Number: 67-63-0, Sigma Aldrich) for 30 minutes, air-dried at 22 oC for 30 minutes, and placed in an oven at 40 oC in humidified air for 48 hours (VO1824HPC, Lindberg/Blue M Vacuum Oven 127.4L, Thermo Scientific, Waltham, MA, USA).
3. To check particle emission pre and post-sterilization, conduct particle count using a particle counter (SOLAIR 3100, Lighthouse Worldwide Solutions), detecting sizes 0.3 to 10 microns, for 1-minute cycles.
4. To check particle emission in humidified air, place part in humidified warm air at 40 oC. Repeat step above to assess particle emission.
Vent-Lock 1+n(1) circuit and components.
1. Acquire Vent-Lock circuit components as assembled in Fig. 1. Vent-Lock 3DP splitters, flow regulator, and manometer adaptors were used. Commercial components include manometer (Ambu Disposable Pressure Manometer, Ambu, Copenhagen, Denmark), one-way valves (22F x 22M, REF 50245, Mallinckrodt Pharmaceuticals), disposable bacteria filters (BSF104, Vincent Medical), and ventilator tubing (SKU: 999027588, Hudson Rci).
2. Adjust ventilator settings. Place the ventilator (Puritan Bennett 840 Ventilator System, Avante Health Solutions) on pressure control mode of ventilation (Volume Ventilation Plus™, Avante Health Solutions) with additional settings detailed in Fig 2.
3. Connect circuit as depicted in Fig 1. to two test lungs (Standard patient: Rp = 2 cmH2O/L/s, RespiTrainer Advance, QuickLung, IngMar Medical; Variable patient: Rp= 50 cmH2O/L/s, ASL 5000, IngMar Medical).
4. Collect intrapulmonary data for both patients: peak inspiratory pressures, tidal volumes, and peak end expiratory pressures. Collect five total values of tidal volume per data set, to be averaged. Also collect corresponding ventilator data, including total expiratory volumes, peak inspiratory pressures, mean inspiratory pressures, and peak end expiratory pressures.
In vivo swine studies.
1. Confirm that in vivo swine studies are compliant with the Guide for the Care and Use of Laboratory Animals, and approved by the Institutional Animal Care and Use Committee by your institution.
2. Acquire two domestic swine (Sus scrofa domesticus, Oak Hill Genetics, Ewing, IL), and allow minimum of seven day acclimation period.
3. Sedate swines with a telazol, ketamine, xylazine cocktail, and intubate with a 7.0 endotracheal tube. Maintain anesthesia with isoflurane.
4. Perform femoral venous and arterial catheterization. Maintain standard ASA monitoring throughout the experiment.
5. Connect the two swines to a single ventilator or anesthesia gas machine (Drager Narkomed 2A) using the Vent-Lock circuit a depicted in Figure 1.
6. Measure flow from each expiatory limb with a SS11LB airflow transducer (Biopac; Goleta, CA). Collect flow data at 2kHz using an MP36 data acquisition unit and BSL 4.1.3 software (Biopac; Goleta, CA).
7. Smooth the spirometry data was then smoothed with a 0.25 sec wide moving median filter after removal of instrument noise below 0.08 L/sec (determined by histogram inspection). Numerically integrate the smoothed data to estimate respiratory tidal volume and use a first order numeric derivative to calculate the instantaneous respiratory rate. Determine the noise floor for the integrated volume by histogram inspection: the resulting threshold should approximate 90 mL.
8. Align the anesthesia record and the spirometry results using common timestamps. Remove all breaths spontaneously initiated by the swine (identified by respiratory rates more than 30% away from the ventilator set point) from analysis.
9. Calculate the mean and standard deviation for each anesthesia record entry for respiratory rate, tidal volume, minute ventilation, and lung compliance.
10. A custom MATLAB script (MATLAB 2019b, The MathWorks, Inc., Natick, MA)] is available for download of all describe analysis.
11. Collect arterial and venous blood gas data every 15 minutes following any changes to the Vent-Lock 3DP device.
12. Following the procedure, euthanize the swine with an overdose (~150mg/kg) of supersaturated potassium chloride IV while under anesthesia.
13. Perform necropsy to assess for any gross lung pathology.