TABLE 2.

Evaluation of staining reagents for cytometer detection

Primary antibody (origin)Type of detection (origina)SampleResult (PIb)Background level
Polyclonal antibodiesGreen
    Lp sg 1 (NRCL)    Rabbit Ig, secondary antibody,c FITC (Dako)Lp sg 1 cells>1,000Low
    Lp sg 1 (NRCL)    Rabbit Ig secondary antibody, HRP, tyramide- FITC (Dako)Lp sg 1 cellsNo staining
    Lp sg 1 HRP (NRCL)    Tyramide-FITC (NEN)Lp sg 1 cellsNo staining
    Lp sg 1 biotin (NRCL)    Streptavidin-FITC (Dako)Lp sg 1 cells450Medium
    Lp FITC (Accurate Chemical & Scientific)Lp sg 1 cells150Low
    Lp FITC (Accurate Chemical & Scientific)    Rabbit Ig secondary antibody, FITC (Dako)Lp sg 1 cells>1,000Low
MAbs
    Monofluo kit Lp sg 1-14 FITC (Bio-Rad)Lp sg 1 cells300Low
    Monofluo kit Lp sg 1-14 FITC (Bio-Rad)    Mouse IgG secondary antibody, FITC (Sigma)Lp sg 1 cells600High
    Lp sg 1-14 Maine Biotechnology Services    Mouse IgG secondary antibody, FITC (Sigma)Lp sg 1 cells250Low
    Lp sg 1-14 (Research Diagnostics)    Mouse IgG secondary antibody, FITC (Sigma)Lp sg 1 cellsNo staining
    Legionella (Dresden)    Mouse IgG secondary antibody, FITC (Sigma)Lp sg 1 cellsNo staining
    Lp sg 1 (Dresden)    Mouse IgG secondary antibody, FITC (Sigma)Lp sg 1 cells400-500Low
    Lp sg 1 (Dresden)    Mouse IgG secondary antibody, FITC (Sigma)Natural water200Medium
    Lp sg 1 (Dresden)    Mouse IgG secondary antibody, FITC, + tertiary antibody, FITC (Sigma, Dako)Lp sg 1 cells900-1,000High
    Lp sg 1 (Dresden)    En Vision + mouse IgG secondary antibody, HRP (Dako)Lp sg 1 cellsNo staining
    Lp sg 2-6, 8-10, 12-15 (Dresden)    Mouse IgG secondary antibody, FITC (Sigma)Lp sg 3 cells300-400Low
    Lp sg 2-6, 8-10, 12-15 (Dresden)    Mouse IgG secondary antibody, FITC (Sigma)Natural water250Medium
    Lp sg 2-6, 8-10, 12-15 (Dresden)    Mouse IgG secondary antibody, FITC, + tertiary antibody, FITC (Sigma, Dako)Lp sg 3 cells750High
Polyclonal antibodiesRed
    Lp sg 1 biotin (NRCL)    Streptavidin-RPE-Cy5 (Dako)Lp sg 1 cells400-500dVery high
    Lp sg 1 biotin (NRCL)    Streptavidin-Alexa Fluor-RPE 647 nm (Molecular probes)Lp sg 1 cells700dVery high
    Lp sg 1 (NRCL)    Rabbit IgG secondary antibody, Alexa Fluor-RPE 647 nm (Molecular Probes)Lp sg 1 cells600dVery high
  • a NEN, Boston, Mass.

  • b PI, mean peak intensity, measured in arbitrary fluorescence units.

  • c Ig, immunoglobulin.

  • d A part of the filtered bacteria was not detected, because their fluorescence was below the limit of detection.