This is Part 2 of a presentation on Genetic Toxicology that was given by Dr. David Kirkland to scientific staff at Health Canada in Nov. 2010. Part 1 is availabile here in ppt and as a webinar at the LinkedIn DABT CE group link

1. David Kirkland

3. DMEM F-10 F-12 RPMI RPMI McCoys Williams Hepes 5A E  M of H 2 O 2 produced - + - + - + - + - + - + - + EGCG Sig. clastogenic dose of H 2 O 2

4. % cells with CA (solid line) Relative cell count (%) (dashed line) Incubation time (hr) of EGCG in medium before stopping reaction McCoy’s 5A F-10

6. Chemical Previous LEC New LEC Allyl isovalerate 2.81 mM in MLA +ve at 0.55 mM in CA (3 hr +S9) Benzofuran 1.27 mM in MLA -ve at in MLA up to 1 mM (64-99% toxicity ). Increased MF at 2 mM but almost 100% toxicity Caffeic acid 1.11 mM in MLA +ve 0.4 mM in MLA (24 hr –S9) and 1 mM in CA (20 hr –S9) Chlorobenzene 1.11 mM in MLA +ve at 0.6 mM in MLA (3 hr +S9) Daminozide 13.75 mM in CA; 11.25 mM in MLA -ve up to 10 mM (CA & MLA) Furan 1.47 mM in CA +ve at 0.8 mM in MLA (3 hr +S9) and at 4 mM in CA (3 hr +S9) Methylolacrylamide 2.94 mM in CA +ve at 2 mM in CA (20 hr –S9) and MLA (24 hr –S9) Toluene 2.44 mM in MLA -ve up to toxic doses (10% RTG) in MLA (not tested in CA) Monuron 6.54 mM in CA -ve in CA up to 4.25 mM which induced >50% toxicity (- and + S9)

8. % cells with CA Survival measure relative to control Conc. of drug(  g/ml) 50% toxicity

10. Negative Positive Positive Equivocal *** p≥0.001

31. In vivo micronucleus assay 6-8 h 18-22 h Erythroblasts Immature erythrocytes Mature erythrocytes Bone marrow Peripheral blood Clastogen

36. Peripheral Blood MN Methodology Inside the Cytometer FITC PI 488nm laser NCE (No Fluorescence) Reticulocyte (Green) MN-Reticulocyte (Green + Red) Platelet (Yellow) Key PE

37. EMS - 3 hr Peripheral Blood MN Methodology Data Output

42. MN % mg/kg

44. MN/1000 PCE mg/kg Drug X

49. Cells with DNA migration Cells without DNA migration

53. Cohesive end BamH1 BamH1 BamH1 BamH1 XHol BamH1 EcoR1 EcoR1 EcoR1 EcoR1 BamH1 Cohesive end LacZ 0 50 kb