In Vitro Binding of 99mTc-Labeled Defensins 1–3 and Ciprofloxacin to Microorganisms and Activated Human Cells
| 99mTc-labeled compound | S. aureus* | K. pneumoniae | C. albicans† | Leukocytes‡ | Endothelial cells§ |
|---|---|---|---|---|---|
| Defensins 1–3 | 48 ± 10 | 37 ± 6 | 17 ± 1 | 5 ± 1 | 3 ± 1 |
| Ciprofloxacin | 5 ± 1 | 6 ± 2 | 6 ± 1 | 17 ± 2 | 20 ± 5 |
Binding of 99mTc-labeled defensins 1–3 and ciprofloxacin to microorganisms was assessed at 4°C as described (1,2). In short, 1 × 107 CFU of microorganisms in 10-mmol/L sodium phosphate buffer (pH, 7.4; NaPB) was added to a mixture containing 99mTc-labeled ciprofloxacin or defensins 1–3 (1 nmol) in NaPB supplemented with 50% v/v 0.01 mol/L acetic acid and 0.01% v/v between −80 (final pH, 5) and incubated for 1 h. Next, microorganisms were washed twice and radioactivity in cell pellet was measured in dose calibrator. Radioactivity associated with microorganisms is expressed as percentage of added radioactivity bound to 2 × 107 microorganisms. Binding of 99mTc-labeled defensins 1–3 and ciprofloxacin to activated human leukocytes or endothelial cells was determined under similar conditions, except that NaPB was supplemented with 50 IU of heparin/mL. Values (means and SEMs) are expressed as percentage of radioactivity bound to 2 × 107 CFU of microorganisms or 2 × 107 human cells. Results are from 6 independent experiments.
↵* Multi–drug-resistant S. aureus type 2141 is a clinical isolate (Dept. of Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands).
↵† Fluconazole-resistant C. albicans Y-01 was obtained from Pfizer Inc., New York, New York.
↵‡ Monocytes and granulocytes were isolated from blood, mixed, and then incubated with 100 nmol/L of synthetic bacterial breakdown product formyl-MLP and 100 ng LPS/mL.
↵§ Human umbilical cord endothelial cells were incubated with 10 U/mL of recombinant IL-1β.