A fixed cytometer chip for identification of cell populations and real-time monitoring of single-cell apoptosis under gradient UV radiation

  • Anyue Xia
  • Mingzhe GanEmail author
  • Huan Xu
  • Yiheng Zhang
  • Dandan Wang
  • Jing Du
  • Qian Sun
  • Jiana Jiang
  • Dan Luo
  • Jinhui CuiEmail author
  • Peifeng LiuEmail author
Research Paper


Cytometry is a basic method to determine cell populations and morphology. Flow cytometry and hemocytometry are the two most common methods among cytometric technologies. However, flow cytometry needs bulky and expensive equipment as well as professional operations, while hemocytometry is limited by its simple function. Both of them are not suitable for real-time monitoring of the morphological changes of single cells. Here, we developed a fixed cytometer chip with two functional modes for both identification of cell populations (I-mode) and real-time monitoring of single-cell morphological changes (M-mode). In I-mode, the fixed cytometer chip was employed to evaluate the cell populations, the results were in accordance with those from the hemocytometer counting and flow cytometry. Besides that, the cell populations were further precisely identified by measuring two-color fluorescence intensities of single cells, which were consistent with the dual parameter analysis in flow cytometry. In M-mode, the chip was applied to real-time monitoring of the single-cell apoptosis under gradient UV radiation, generated by a novel stair-like UV shield. The dynamic apoptotic morphologies of a large number of single cells were monitored in real-time by time-lapse imaging. In addition, we integrated eight parallel channels on a 60 mm × 30 mm chip, and the chip could achieve scalable single-cell capture and analysis capability. This fixed cytometer chip is bifunctional, easy-to-handle, universal, and scalable.


Cytometer chip Cell population identification Real-time monitoring Cell Apoptosis Gradient UV radiation 



We gratefully acknowledge the financial support from the National Natural Science Foundation of China (81771968, 81472842, 21778071, and 31400087), Shanghai Municipal Education Commission-Gaofeng Clinical Medicine Grant Support (20181705), Shanghai Talent Development Fund (2017053), Translational Medicine Cross Research Grant of Shanghai Jiao Tong University (ZH2018ZDA05), Youth Innovation Promotion Association CAS and Suzhou Institute of Nano-Tech and Nano-Bionics Owned Fund (Y5AAS11001).

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

10404_2019_2244_MOESM1_ESM.docx (1.3 mb)
Supplementary material 1 (DOCX 1345 kb)


  1. Beck M, Brockhuis S, van der Velde N, Breukers C, Greve J, Terstappen LW (2012) On-chip sample preparation by controlled release of antibodies for simple CD4 counting. Lab Chip 12:167–173. CrossRefGoogle Scholar
  2. Byrne SN, Kok L, Leighton S, Marsh-Wakefield F, Halliday GM (2013) UV-induced immune suppression: implications for autoimmune disease. Australas J Dermatol 54:28. CrossRefGoogle Scholar
  3. Cheng YH, Chen YC, Brien R, Yoon E (2016) Scaling and automation of a high-throughput single-cell-derived tumor sphere assay chip. Lab Chip 16:3708–3717. CrossRefGoogle Scholar
  4. Cho SH, Chen CH, Tsai FS, Godin JM, Lo YH (2010) Human mammalian cell sorting using a highly integrated micro-fabricated fluorescence-activated cell sorter (microFACS). Lab Chip 10:1567–1573. CrossRefGoogle Scholar
  5. Cui X, Yip HM, Zhu Q, Yang CP, Lam RHW (2014) Microfluidic long-term differential oxygenation for bacterial growth characteristics analyses. Rsc Adv 4:16662–16673. CrossRefGoogle Scholar
  6. Del Carratore R, Della Croce C, Simili M, Taccini E, Scavuzzo M, Sbrana S (2002) Cell cycle and morphological alterations as indicative of apoptosis promoted by UV irradiation in S-cerevisiae. Mut Res Gen Toxicol Environ 513:183–191. CrossRefGoogle Scholar
  7. Deshmukh J, Pofahl R, Haase I (2017) Epidermal Rac1 regulates the DNA damage response and protects from UV-light-induced keratinocyte apoptosis and skin carcinogenesis. Cell Death Dis 8:e2663. CrossRefGoogle Scholar
  8. Di Carlo D, Aghdam N, Lee LP (2006) Single-cell enzyme concentrations, kinetics, and inhibition analysis using high-density hydrodynamic cell isolation arrays. Anal Chem 78:4925–4930. CrossRefGoogle Scholar
  9. Douek DC et al (2002) HIV preferentially infects HIV-specific CD4(+) T cells. Nature 417:95–98. CrossRefGoogle Scholar
  10. Durham ND, Chen BK (2016) Measuring T cell-to-T cell HIV-1 transfer, viral fusion, and infection using flow cytometry methods in molecular biology. Clifton NJ 1354:21–38. CrossRefGoogle Scholar
  11. Eom H-J, Choi J (2010) p38 MAPK Activation, DNA damage, cell cycle arrest and apoptosis as mechanisms of toxicity of silver nanoparticles in Jurkat T cells. Environ Sci Technol 44:8337–8342. CrossRefGoogle Scholar
  12. Fan YM, Wu DM, Jin LN, Yin ZM (2005) Human glutamylcysteine synthetase protects HEK293 cells against UV-induced cell death through inhibition of c-Jun NH2-terminal kinase. Cell Biol Int 29:695–702. CrossRefGoogle Scholar
  13. Gao D, Li H-F, Guo G-S, Lin J-M (2010) Magnetic bead based immunoassay for enumeration of CD4(+) T lymphocytes on a microfluidic device. Talanta 82:528–533. CrossRefGoogle Scholar
  14. Gervais O, Renault T, Arzul I (2015) Induction of apoptosis by UV in the flat oyster, Ostrea edulis Fish Shellfish. Immun 46:232–242. CrossRefGoogle Scholar
  15. Grossman Z, Meier-Schellersheim M, Sousa AE, Victorino RM, Paul WE (2002) CD4 + T-cell depletion in HIV infection: are we closer to understanding the cause? Nat Med 8:319–323. CrossRefGoogle Scholar
  16. Guo Q, Duffy SP, Matthews K, Islamzada E, Ma HS (2017) Deformability based cell sorting using microfluidic ratchets enabling phenotypic separation of leukocytes directly from whole blood. Sci Rep 7:6627. CrossRefGoogle Scholar
  17. Huang C-H, Hou H-S, Lo K-Y, Cheng J-Y, Sun Y-S (2017) Use microfluidic chips to study the effects of ultraviolet lights on human fibroblasts. Microfluid Nanofluid 21:79. CrossRefGoogle Scholar
  18. Kemeny L, Csoma Z, Bagdi E, Banham AH, Krenacs L, Koreck A (2010) Targeted phototherapy of plaque-type psoriasis using ultraviolet B-light-emitting diodes. Brit J Dermatol 163:167–173. CrossRefGoogle Scholar
  19. Kimura H et al (2010) UV light killing efficacy of fluorescent protein-expressing cancer cells in vitro and in vivo. J Cell Biochem 110:1439–1446. CrossRefGoogle Scholar
  20. Liang L, Jin YX, Zhu XQ, Zhou FL, Yang Y (2018) Real-time detection and monitoring of the drug resistance of single myeloid leukemia cells by diffused total internal reflection. Lab Chip 18:1422–1429. CrossRefGoogle Scholar
  21. Lin CH et al (2015) A microfluidic dual-well device for high-throughput single-cell capture and culture. Lab Chip 15:2928–2938. CrossRefGoogle Scholar
  22. Luo Q et al (2018) Overexpression of CD64 on CD14(++)CD16(-) and CD14(++)CD16(+) monocytes of rheumatoid arthritis patients correlates with disease activity. Exp Ther Med 16:2703–2711. CrossRefGoogle Scholar
  23. Noor AM et al (2018) A microfluidic chip for capturing, imaging and counting CD3(+) T-lymphocytes and CD19(+) B-lymphocytes from whole blood. Sens Actuators B Chem 276:107–113. CrossRefGoogle Scholar
  24. O’Neill K, Aghaeepour N, Spidlen J, Brinkman R (2013) Flow cytometry bioinformatics. Plos Comput Biol 9:e1003365. CrossRefGoogle Scholar
  25. Pierzchalski A, Mittag A, Tarnok A (2011) Introduction a: recent advances in cytometry instrumentation, probes, and methods-review. In: Darzynkiewicz Z, Holden E, Orfao A, Telford W, Wlodkowic D (eds) Recent advances in cytometry, part a: instrumentation, methods, Fifth Edition, vol 102. Methods in Cell Biology. pp 1–21. Google Scholar
  26. Piruska A, Nikcevic I, Lee SH, Ahn C, Heineman WR, Limbach PA, Seliskar CJ (2005) The autofluorescence of plastic materials and chips measured under laser irradiation. Lab Chip 5:1348–1354. CrossRefGoogle Scholar
  27. Polinkovsky M, Gutierrez E, Levchenko A, Groisman A (2009) Fine temporal control of the medium gas content and acidity and on-chip generation of series of oxygen concentrations for cell cultures. Lab Chip 9:1073–1084. CrossRefGoogle Scholar
  28. Racz E, Prens EP (2015) Phototherapy and photochemotherapy for psoriasis. Dermatol Clin 33:79–89. CrossRefGoogle Scholar
  29. Rawstron AC, de Tute RM, Haughton J, Owen RG (2016) Measuring disease levels in myeloma using flow cytometry in combination with other laboratory techniques: lessons from the past 20 years at the leeds haematological malignancy diagnostic service. Cytom Part B Clin Cytom 90:54–60. CrossRefGoogle Scholar
  30. Rettig JR, Folch A (2005) Large-scale single-cell trapping and imaging using microwell arrays. Anal Chem 77:5628–5634. CrossRefGoogle Scholar
  31. Salmhofer W, Soyer HP, Wolf P, Fodinger D, Hodl S, Kerl H (2004) UV light-induced linear IgA dermatosis. J Am Acad Dermatol 50:109–115. CrossRefGoogle Scholar
  32. Sarkar S, Rajput S, Tripathi AK, Mandal M (2013) Targeted therapy against EGFR and VEGFR using ZD6474 enhances the therapeutic potential of UV-B phototherapy in breast cancer cells. Mol Cancer 12:122. CrossRefGoogle Scholar
  33. Seidi A, Kaji H, Annabi N, Ostrovidov S, Ramalingam M, Khademhosseini A (2011) A microfluidic-based neurotoxin concentration gradient for the generation of an in vitro model of Parkinson’s disease. Biomicrofluidics 5:022214. CrossRefGoogle Scholar
  34. Shields CW, Reyes CD, Lopez GP (2015) Microfluidic cell sorting: a review of the advances in the separation of cells from debulking to rare cell isolation. Lab Chip 15:1230–1249. CrossRefGoogle Scholar
  35. Skelley AM, Kirak O, Suh H, Jaenisch R, Voldman J (2009) Microfluidic control of cell pairing and fusion. Nat Methods 6:147–152. CrossRefGoogle Scholar
  36. Suh S-S, Oh SK, Lee SG, Kim I-C, Kim S (2017) Porphyra-334, a mycosporine-like amino acid, attenuates UV-induced apoptosis in HaCaT cells. Acta Pharmaceut 67:257–264. CrossRefGoogle Scholar
  37. Tai MH, Weng CH, Mon DP, Hu CY, Wu MH (2012) Ultraviolet C irradiation induces different expression of cyclooxygenase 2 in NIH 3T3 cells and A431 cells: the roles of COX-2 are different in various cell lines. Int J Mol Sci 13:4351–4366. CrossRefGoogle Scholar
  38. Tan WH, Takeuchi S (2007) A trap-and-release integrated microfluidic system for dynamic microarray applications. Proc Natl Acad Sci U S A 104:1146–1151. CrossRefGoogle Scholar
  39. Tang W, Tang D, Ni Z, Xiang N, Yi H (2017) Microfluidic impedance cytometer with inertial focusing and liquid electrodes for high-throughput cell counting and discrimination. Anal Chem 89:3154–3161. CrossRefGoogle Scholar
  40. Voos P et al (2018) Ionizing radiation induces morphological changes and immunological modulation of Jurkat cells. Front Immunol 9:922. CrossRefGoogle Scholar
  41. Wang X, Yang C, Xu F, Qi L, Wang J, Yang P (2018) Imbalance of circulating Tfr/Tfh ratio in patients with rheumatoid arthritis. Clin Exp Med 19:1–10. CrossRefGoogle Scholar
  42. Wasserberg D et al (2018) All-printed cell counting chambers with on-chip sample preparation for point-of-care CD4 counting. Biosens Bioelectron 117:659–668. CrossRefGoogle Scholar
  43. Weng L et al (2017) A highly-occupied, single-cell trapping microarray for determination of cell membrane permeability. Lab Chip 17:4077–4088. CrossRefGoogle Scholar
  44. Wlodkowic D, Faley S, Zagnoni M, Wikswo JP, Cooper JM (2009) Microfluidic single-cell array cytometry for the analysis of tumor apoptosis. Anal Chem 81:5517–5523. CrossRefGoogle Scholar
  45. Xu GG, Marcusson JA, Hemminki K (2001) DNA photodamage induced by UV phototherapy lamps and sunlamps in human skin in situ and its potential importance for skin cancer. J Invest Dermatol 116:194–195. CrossRefGoogle Scholar
  46. Xu J et al (2018) A microfluidic chip with double-slit arrays for enhanced capture of single cells. Micromachines-Basel 9:157. CrossRefGoogle Scholar
  47. Yao J et al (2013) Ultraviolet (UV) and hydrogen peroxide activate ceramide-ER stress-AMPK signaling axis to promote retinal pigment epithelium (RPE) cell apoptosis. Int J Mol Sci 14:10355–10368. CrossRefGoogle Scholar
  48. Yun H, Bang H, Min J, Chung C, Chang JK, Han DC (2010) Simultaneous counting of two subsets of leukocytes using fluorescent silica nanoparticles in a sheathless microchip flow cytometer. Lab Chip 10:3243–3254. CrossRefGoogle Scholar
  49. Zhang Y et al (2017) Detection of sepsis in patient blood samples using CD64 expression in a microfluidic cell separation device. Analyst 143:241–249. CrossRefGoogle Scholar
  50. Zhu H, Macal M, Jones CN, George MD, Dandekar S, Revzin A (2008) A miniature cytometry platform for capture and characterization of T-lymphocytes from human blood. Anal Chim Acta 608:186–196. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Central Laboratory, Renji Hospital, School of MedicineShanghai Jiao Tong UniversityShanghaiChina
  2. 2.State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, School of MedicineShanghai Jiao Tong UniversityShanghaiChina
  3. 3.Micro-Nano Research and Diagnosis Center, Renji Hospital, School of MedicineShanghai Jiao Tong UniversityShanghaiChina
  4. 4.CAS Key Laboratory of Nano-Bio Interface, Suzhou Institute of Nano-Tech and Nano-BionicsChinese Academy of SciencesSuzhouChina
  5. 5.Department of Clinical Laboratory Medicine, Southwest HospitalArmy Medical University (Third Military Medical University)ChongqingChina
  6. 6.Kavli Institute at Cornell for Nanoscale ScienceCornell UniversityIthacaUSA
  7. 7.Department of Biological and Environmental EngineeringCornell UniversityIthacaUSA

Personalised recommendations