Immunogold Labeling of Rubisco In C4 Plant Leaves for Scanning Electron Microscopy

Abstract

Immunogold labeling was applied to scanning electron microscopy to examine the inner structure of leaves of G4 plants, maize and linger millet. Ultrastructural features of the G4 plant leaves were observed on cross sections in the secondary electron imaging mode of the scanning electron microscope. Immunogold localization was well recognized in the backscattered imaging mode at the low accelerating voltage of 5 kV on the specimens treated with tannic acid and Oso4 instead of metal coating. Gold labeling of ribulose 1,5-bisphosphate carboxylase/oxygenase was specifically distributed on the cut surface of bundle sheath chloroplasts of the C4 plants. Gold particles were scarcely observed in mesophyll chloroplasts or other organelles. Freeze-cracking of the leaves was found to be effective to expose the inner structure of the organelles. The secondary and backscattered electron images were also compared under various accelerating voltages.

Keywords

• Backscattered electron
• Bundle sheath chloroplast
• C4 plant
• Immunogold localization
• Ribulose 1,5-bisphosphate carboxylase/oxygenase
• Scanning electron microscopy
• Secondary electron