As a new generation of polyethylene imine transfection reagent, PEIpro® has basically replaced the early linear/branched chain polyethylene imine and other similar products, and has been widely used in the r&d and production of vaccines, CAR-T therapy and gene therapy products.  Because the transfection reagent has certain cytotoxicity and is not easy to degrade, it is of great significance to develop its own assay method.  However, due to intellectual property rights, technical protection and other factors, the content detection methods of PEIpro® have not been publicly reported, and the residue detection work of PEIpro® can only rely on a few European companies.

In order to solve the detection problem of this transfection reagent and better serve local pharmaceutical enterprises, Shanghai Xihua Pharmaceutical Co., Ltd. and Shimadzu Company jointly developed the exclusive detection method for PEIpro®.  The shim-Pack Scepter-HD-C18 (150*4.6mm, 3μm) column was found to be suitable for the detection of the transfection reagent by screening a large number of columns.  As shown in Figure 1, this column has very good retention of PEIpro and good peak form.  

Figure 1   Chromatogram of 0.5μg/ ml spiked solution of PEIpro

1、Precision of peak area (n=6)

The control solution was injected for 6 times, and the results (see Figure 2) showed good reproducibility and precision of the method.

Figure 2   6 needle stacking diagram of control solution

2、Detection sensitivity

As shown in Figure 3, the limits of detection and quantification of this method can be calculated to be about 0.003μg /mL and 0.01μg /mL, indicating that the method has extremely high detection sensitivity.

Figure 3. 0.05 and 0.15μg/ mL PEIpro control solutions

3、Standard work curve

The results are shown in Figure 4, R2=1.00, and the fitting results are excellent.

Figure 4 Working curve of 0.05μg/ml-1.0μg/ mL concentration range

4、Method accuracy

As shown in FIG. 5, this method has excellent accuracy and is less disturbed by the substrate of biological samples.

Figure 5 Comparison of 0.5μg/ mL control solution and 0.5μg/ mL standard solution

5、Summary

In summary, this method has good accuracy and precision, excellent correlation of working curve, high sensitivity and strong anti-interference ability, which can be used as the preferred method for PEIpro® residue detection.