TABLE 1.

Relative fluorine and bromine abundance of E. coli and V. cholerae cells after FISH

Method, probe, and/or dye (n)aNo. of single cells analyzedRelative halogen abundance of a set of single cellsfSignal/background ratio
19F/12C19F/14N81Br/12C81Br/14NHalogen/ CHalogen/ N
Standard FISH with halogen-containing probe EUB338-I with Cy3 and 5Fl-dUb164.5 × 10−3 ± 1.3 × 10−45.0 × 10−3 ± 1.2 × 10−426
EL-FISH with different tyramides on E. colic
    517F74.1 × 10−1 ± 1.4 × 10−22.3 × 10−1 ± 4.2 × 10−3181265
    570F61.8 × 10−2 ± 4.6 × 10−41.1 × 10−2 ± 2.1 × 10−4813
    544Br111.8 × 10−2 ± 1.4 × 10−46.5 × 10−3 ± 8.7 × 10−52221
    Background controld2.3 × 10−3 ± 7.0 × 10−58.8 × 10−4 ± 4.2 × 10−58.1 × 10−4 ± 2.1 × 10−53.1 × 10−4 ± 1.3 × 10−511
EL-FISH on Vibrio sp.e
    EUB338-I228.4 × 10−1 ± 1.8 × 10−22.2 × 10−1 ± 6.0 × 10−34237
    NON33892.0 × 10−2 ± 9.0 × 10−46.0 × 10−3 ± 2.0 × 10−311
  • a Standard FISH, fluorine-labeling of E. coli cells with directly fluorine-labeled oligonucleotide probes; EL-FISH on E. coli, comparison of different halogen-containing tyramides after EL-FISH on E. coli cells with probe EUB338-I; EL-FISH on Vibrio sp., comparison of positive (probe EUB338-I) and negative (probe NON338) control hybridizations on V. cholerae cells.

  • b 5Fl-dU nucleotides were substituted for deoxythymidine nucleotides. The EUB338-I probe sequence was [Cy3]GC[5Fl-dU]GCC[5Fl-dU]CCCG[5Fl-dU]AGGAGT. The probe was Cy3 labeled for hybridization control by epifluorescence microscopy.

  • c All hybridizations performed with HRP-conjugated EUB338-I probe. See Materials and Methods section for tyramide abbreviations. Tyramide signal amplification was for 10 min at 46°C with a 1:500 dilution of tyramide stock.

  • d Negative control. Analysis was performed on fixed cells without hybridization and tyramide signal amplification.

  • e Hybridizations performed with HRP-conjugated EUB338-I or NON338 probe. Tyramide signal amplification using tyramide Oregon Green 488-X (517 F). Mass images provided in supplemental data.

  • f Numbers given in the table are average values with standard errors for the number of single cells analyzed.