Tracheal aspirate samples were collected within 72 hours of endotracheal intubation with an inline Ballard suction catheter (Halyard, Alpharetta, GA) connected to a sterile Lukens trap (Covidien, Walpole, MA) using up to 5 mL of sterile saline and processed according to published protocols (19 (link), 20 (link)). For infants and toddlers, 1–2 mL of saline is instilled. The Ballard inline suction catheter is passed once to obtain a sample. If 50% of the instilled volume is not returned, then a repeat passage of the Ballard is performed. The Ballard suction catheter (Halyard, Alpharetta, GA) may be flushed with additional saline to move the aspirate into the Lukens trap (Covidien, Walpole, MA). Cell viability was determined with trypan blue exclusion on a Countess hemocytometer. Cell purity was assessed by Shandon Kwik-Diff (Thermo Scientific, Waltham, MA) staining of cytospin cell preparations (20 (link)). Up to 1 × 106 cells were stored in RNALater at –80°C until RNA was extracted for gene expression analysis.
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Tracheal Aspirate Sampling for Gene Expression
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Catheter
Cell viability
Cells
Endotracheal intubation
Gene expression analysis
Infants
Saline
Sterile
Suction
Tracheal
Trypan blue
Corresponding Organization :
Other organizations : Emory University, Children's Healthcare of Atlanta, Georgia Institute of Technology
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Variable analysis
independent variables
- Tracheal aspirate sample collection within 72 hours of endotracheal intubation
- Cell viability determined with trypan blue exclusion on a Countess hemocytometer
- Cell purity assessed by Shandon Kwik-Diff staining of cytospin cell preparations
- Tracheal aspirate samples collected using an inline Ballard suction catheter (Halyard, Alpharetta, GA) connected to a sterile Lukens trap (Covidien, Walpole, MA)
- Up to 5 mL of sterile saline used to process the samples according to published protocols
- For infants and toddlers, 1–2 mL of saline is instilled
- The Ballard inline suction catheter is passed once to obtain a sample, and if 50% of the instilled volume is not returned, a repeat passage is performed
- The Ballard suction catheter may be flushed with additional saline to move the aspirate into the Lukens trap
- Up to 1 × 10^6 cells stored in RNALater at –80°C until RNA was extracted for gene expression analysis
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