Table 1. Troubleshooting guide.

Step	Problem observed	Possible reason	Solutions
2.3.2	No transduced cells after puromycin selection	Low efficiency of lentiviral transduction	Quantify viral titer before target cell transduction; Optimize the dilution ratio
3.3.11	Fusion construct is undetectable by western blots	Incorrect protocol; Low expression level of the construct	Use a positive control; Optimize construct expression level
3.3.11	High background signal in omit-biotin control group	Excessive non-specific protein binding to streptavidin-coated beads	Increase the number of wash steps; Reduce the amount of streptavidin beads; Shorten incubation time
4.2.3	Cells detach during PBS washing	Poor cell adhesion	Optimize poly-L-lysine concentration and incubation time
4.3.3	GAPDH detected in digitonin-permeabilized groups	Residual cytoplasmic components	Use freshly prepared digitonin solution; Increase the number of wash steps
4.3.3	Low yield of biotinylated proteins	Insufficient samples or streptavidin beads used	Increase the number of cells; Increase the amount of streptavidin beads for protein enrichment