Table 1.
Sample types commonly used for ARDS metabolomics studies
| Sample Type | Preparation | Advantages | Disadvantages | Recommendations |
|---|---|---|---|---|
| BALF/mini-BALF | • Centrifuge to remove cells and debris (800 g at 4°C for 10 min) • Remove supernatant • Aliquot and freeze (−80°C) until the time of assay |
• Collected from specific area of the lung (BALF only) • Collected under clinical and reproducible conditions |
• Invasive and not well tolerated in patients with severe ARDS • Diluted (typically 100X); thus not all metabolites can be assessed • Difficult to accurately normalize for sample dilution • Concentration process is often required leading to variability |
• First-cycle lavage is preferred • Correct for dilution with urea ratio (e.g., BALF-plasma) • Consider buffer exchange to remove salt and methanol or acetone precipitation for protein separation |
| • Concentration process is often required leading to variability | ||||
| • Cannot be used easily for longitudinal sampling | ||||
| • Mini-BALF is less standardized compared with BALF | ||||
| Exhaled breath condensate | • Collect during tidal breathing using a nose clip and a saliva trap • Define cooling temperature and collection time • Use inert material for condenser • Do not use resistor and do not use filter between the subject and the condenser |
• Noninvasive • Safe • Suitable for analysis of nonvolatile components • Suitable for longitudinal study • Feasible in children |
• Very diluted • Difficult to standardize • Often requires concentration steps leading to variability • Potential variability due to differences in droplet dilution • Samples whole airway; difficult to localize changes |
• Consider commercial equipment, such as EcoScreen or RTube • Collection time of 10 min generally sufficient to obtain 1–2 ml of sample and is well tolerated by patients |
| • Difficult to normalize metabolites for the total content | ||||
| • High variability in sample quality | ||||
| Plasma/serum | • Collect blood by direct venipuncture, if possible, into a Vacutainer tube | |||
| • For plasma, make sure the Vacutainer tube contains either EDTA or sodium heparin; immediately invert the tube several times to ensure mixture with anticoagulant • For serum, make sure the Vacutainer tube has no additive; allow the blood to clot at room temperature for at least 30 min • After 30 min of blood collection, centrifuge balanced tubes (15 min at 1,300 g) with no brake to ensure proper separation |
• Minimally invasive • Easy to collect with standardized protocols • Widely used in metabolomics studies • Composition relatively well documented • Relatively consistent and easy to define protocol in multicenter studies • Contains a large number of potential targets |
• Plasma is not well suited for NMR especially if filters are used • Lipid composition dominated by lipoproteins, possibly masking minor components • Distant from the tissue of interest, so there is potential bias toward systemic changes in disease |
• Refrigeration before or during plasma centrifugation is recommended | |
| • After centrifugation, use the upper layer (clear and pale yellow in color) and avoid disturbing other layer(s) | ||||
| • Carefully aliquot and freeze (−80°C) in Cryovial |