• Determine the in vivo ratio of tubulin heterodimer to microtubules in cell or tissue lysates
• Study the effects of pharmaceutical compounds on the ratio of tubulin to microtubules

Kit content (sufficient to process 100 x 1 ml lysates)

• Lysis & microtubule stabilization buffer
• Protease inhibitor cocktail
• ATP
• GTP
• Microtubule depolymerizing buffer
• Microtubule-enhancing solution
• Tubulin protein standard
• Anti-tubulin antibody (sheep)
• Anti-sheep secondary HRP conjugated antibody
• SDS sample buffer
• DMSO

Equipment & materials required

• Western blot assay & detection equipment
• Ultracentrifuge: 100,000g, 37°C, 100-1000 µl volume tube capacity
• Small volume homogenizer (low ml)

This kit allows users to measure the ratio of microtubule to free tubulin in cells. Cells or tissues are lysed in a stabilization buffer that preserves the native microtubule-to-tubulin ratio. After lysis, centrifugation separates polymerized microtubules (pellet) from unpolymerized tubulin (supernatant). Tubulin in each fraction is then quantified via western blotting, providing a clear measure of microtubule versus free tubulin content.

Key characteristics

• Semi-quantitative: accurately measures the in vivo microtubule:tubulin ratio
• Separation process: simple ultracentrifugation step
• Detection method: SDS-PAGE or Western blot
• Timeframe: Approximately 8h, typically performed over two days