Dilution cloning

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Dilution cloning or cloning by limiting dilution [1][2] describes a procedure to obtain a monoclonal cell population starting from a polyclonal mass of cells. This is achieved by setting up a series of increasing dilutions of the parent (polyclonal) cell culture. A suspension of the parent cells is made. Appropriate dilutions are then made, depending on cell number in the starting population, as well as the viability and characteristics of the cells being cloned. After the final dilutions are produced, aliquots of the suspension are plated[1] or placed in wells[2] and incubated. If all works correctly, a monoclonal cell colony will be produced. Applications for the procedure include cloning of parasites,[3] T cells,[4] transgenic cells,[5] and macrophages.[6]


  1. ^ a b Freshney, R. Ian (2010). Culture of animal cells : a manual of basic technique and specialized applications (6th ed.). Hoboken, N.J.: Wiley-Blackwell. pp. 208–211. ISBN 9780470528129.
  2. ^ a b Davis, edited by John M (2011). Animal cell culture. Oxford: Wiley-Blackwell. pp. 239–240. ISBN 978047066658-6.
  3. ^ Butterworth, Alice S; Robertson, Alan J; Ho, Mei-Fong; Gatton, Michelle L; McCarthy, James S; Trenholme, Katharine R (2011). "An improved method for undertaking limiting dilution assays for in vitro cloning of Plasmodium falciparum parasites". Malaria Journal. 10 (1): 95. doi:10.1186/1475-2875-10-95. ISSN 1475-2875.
  4. ^ Rout, Namita; Else, James G.; Yue, Simon; Connole, Michelle; Exley, Mark A.; Kaur, Amitinder (2 July 2010). "Heterogeneity in phenotype and function of CD8+ and CD4/CD8 double-negative Natural Killer T cell subsets in sooty mangabeys". Journal of Medical Primatology. 39 (4): 224–234. doi:10.1111/j.1600-0684.2010.00431.x. PMC 2904642.
  5. ^ Lilienfeld, Benjamin G.; Crew, Mark D.; Forte, Pietro; Baumann, Bettina C.; Seebach, Jörg D. (1 March 2007). "Transgenic expression of HLA-E single chain trimer protects porcine endothelial cells against human natural killer cell-mediated cytotoxicity". Xenotransplantation. 14 (2): 126–134. doi:10.1111/j.1399-3089.2007.00378.x.
  6. ^ Seki, Yoshihiro; Suzuki, Satoshi O.; Masui, Kenta; Harada, Shiori; Nakamura, Seiji; Kanba, Shigenobu; Iwaki, Toru (1 June 2011). "A simple and high-yield method for preparation of rat microglial cultures utilizing Aclar plastic film". Neuropathology. 31 (3): 215–222. doi:10.1111/j.1440-1789.2010.01163.x.

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