title: Validation of the flush method as an alternative to basal or reflex tear collection.
abstract: OBJECTIVE
  To validate the more easily applicable "flush" tear collection technique as a viable alternative to basal and reflex tear collection.
  
  
  METHODS
  Total protein content (TPC) and immunoglobulin A (IgA) concentrations were determined in the basal, reflex, and flush tears of 16 healthy non-contact lens wearers. The overall protein profile was established using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry (MS).
  
  
  RESULTS
  Collection-rates were 4.6 ± 6.7 μl/min, 13.9 ± 11.1 μl/min, and 25.7 ± 12.4 μl/min for the basal, reflex, and flush tears, respectively. TPC was 7.14 ± 2.22 mg/mL, 6.01 ± 2.11 mg/mL, and 3.79 ± 1.51 mg/ mL for basal, reflex, and flush tears, respectively, with flush tears being significantly less concentrated than basal (p = 0.001) and reflex (p = 0.008). IgA concentration was 1.04 ± 0.29 mg/ mL, 0.64 ± 0.26 mg/mL, and 0.65 ± 0.23 mg/mL for basal, reflex, and flush tears, respectively, with basal tears being significantly more concentrated (p < 0.001). As a percentage of TPC, IgA represented 19.8 ± 14.9%, 11.4 ± 3.9%, and 19.8 ± 8.7% for basal, reflex, and flush, respectively. The flush was not significantly different to basal (p = 1.00) but significantly greater than reflex (p = 0.02). SDS-PAGE showed the same tear profiles for basal and flush tears. MS identified the most abundant proteins in all tear types.
  
  
  CONCLUSIONS
  The flush method allows much faster collection than basal secretion sampling but returns essentially the same spectrum of proteins in similar proportions. This behavior is confirmation that the flush technique has utility as a more convenient alternative to basal tear sampling in studies involving composition analysis.
authors.affiliation: Brien Holden Vision Institute, Sydney, Australia. m.markoulli@brienholdenvision.org
authors.forename: Maria
authors.initials: M
authors.lastname: Markoulli
authors.forename: Eric
authors.initials: E
authors.lastname: Papas
authors.forename: Andrea
authors.initials: A
authors.lastname: Petznick
authors.forename: Brien
authors.initials: B
authors.lastname: Holden
mesh_terms: D000293:Adolescent; D000328:Adult; D001767:Blinking / Q000502:physiology*; D004591:Electrophoresis, Polyacrylamide Gel; D005136:Eye Proteins / Q000032:analysis*; D005260:Female; D006801:Humans; D007071:Immunoglobulin A, Secretory / Q000032:analysis*; D008297:Male; D013058:Mass Spectrometry; D008875:Middle Aged; D009885:Ophthalmology / Q000379:methods*; D013048:Specimen Handling / Q000379:methods*; D013666:Tears / Q000737:chemistry*; D055815:Young Adult
chemical_list: D005136:Eye Proteins; D007071:Immunoglobulin A, Secretory; C042434:tear proteins