Liver Cancer Cells In Laser Tweezers Raman Spectroscopy

Objective:To study the normal liver cells (L02), the hepatoma cells (HepG2), as well as the fresh tumor tissues after surgery, the adjacent normal tissues (less than 2cm from the surgical margins), normal liver tissue (2cm away from the surgical margins, and the cancer cells can not be found at cut edges) in raman spectra and to establish the relationship between molecular biology, the characteristics of liver cells and Raman spectral library.Method:1.Take logarithmic growth of artificial cultured normal liver cells (L02), hepatoma cells (HepG2), the 5th generations of them were hit scan spectrum;2.Take the patients'confirmed fresh liver specimens(normal and adjacent normal tissue) after surgery. Single cell suspension was prepared by mechanical dispersion, then the single cell suspension was hit Raman spectrometry;3.The single-cell suspension was centrifuged for 15 minutes, and then the background spectrum of the supernatant was collected using the same excitation power and exposure time;4.The final raman spectroscopy of the single cells was got after the raman spectra of single-cell suspension minusing the background spectrum,;5.The origin6.0 and the Unscrambler softwares were used for data processing and PCA analysis. Results:1.The average raman spectra between cultured HepG2,L02 and the raman spectra between generations of HepG2 in the 624 cm-1,853 cm-1,938 cm-1,1001 cm-1,1158 cm-1,1177 cm-1,1259 cm-1,1654 cm-1 (representative of the protein bands),787 cm-1,1094 cm-1 (representative of nucleic acid bands),1309 cm-1, 1448cm-1 (representative of lipid bands),596cm-1,740 cm-1 (band representative of carbohydrates) and band shifts were significantly different; PCA analysis can accurately distinguish the generations of L02, HepG2and HepG2;2.There was no significant difference between the average spectrum of generations of L02;3.. the average spectra of single cells of the cancer tissues, the adjacent normal tissues and the normal tissues in the 621 cm-1,853 cm-1,938 cm-1,1005 cm-1,1158 cm-1,1177 cm-1,1259 cm-1,1654cm-1 (representative of the protein bands),787cm-1,1094cm-1 (representative of nucleic acid bands),718 cm-1, 1448 cm-1 (representative of lipid bands),596 cm-1 (band representative of carbohydrates) peaks have significant differences; PCA analysis can accurately distinguish three types of organizations.Conclusion:1.The characteristic peaks of the cultured HepG2 and L02 were as follows:â‘ characteristic protein peaks were the 624 cm-1,853 cm-1,938 cm-1,1005 cm-1, 1158 cm-1,1177 cm-1,1259 cm-1,1654 cm-1;â‘¡nucleic acid molecules characteristic peaks were the 787 cm-1,1094 cm-1;â‘¢lipid molecules characteristic peaks were the 1309 cm-1,1448 cm-1;â‘£characteristic spectrum of carbohydrate molecules peaks were 596 cm-1,740 cm-1;2.Representative bands of proteins, nucleic acids, lipids, carbohydrates in HepG2 and L02 were significantly different. Raman spectroscopy can accurately distinguish the HepG2 and L02.The individual differences in HepG2 is stronger than that in the L02, while the internal material uniformity was weaker;3. The raman spectra of proteins, nucleic acids, lipids, carbohydrates between the generations of HepG2 (in addition to 5-and 1,3-generationas) epresentative had different bands. Raman spectroscopy can accurately distinguish the generations of HepG2, In its growth process, the cells with the four substances in terms of space and volume were in a very unstable state;4.Representative bands of proteins, nucleic acids, lipids, carbohydrates in generations of L02 showed no significant difference;5.Characteristics peaks of single cells in tissues were as follows:â‘ characteristic protein peaks were the 621 cm-1,853 cm-1,938 cm-1,1005 cm-1, 1158 cm-1,1177 cm-1,1259 cm-1,1654 cm-1;â‘¡nucleic acid molecules characteristic peaks were the 787 cm-1,1094 cm-1;â‘¢lipid molecules characteristic spectral peaks were at 718 cm-1,1448 cm-1;â‘£characteristic spectrum of carbohydrate molecules peaks were the596 cm-1;6.In the liver cancer tissues, adjacent normal tissues and normal tissues,the single-cell proteins, nucleic acids, lipids and carbohydrates had significant difference.Raman spectra can accurately distinguish the single cell of the liver cancer tissues, adjacent normal tissues and normal tissues, which indicated that in the process of the liver carcinogenesis, the spatial structures and the numbers of these above four substances were changed, whit the intracellular destruction of material stability.