Improved Plasmodium falciparum dilution cloning through efficient quantification of parasite numbers and c-SNARF detection
Background Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth i...
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| Published in | Malaria journal Vol. 20; no. 1; pp. 1 - 7 |
|---|---|
| Main Authors | , , |
| Format | Journal Article |
| Language | English |
| Published |
London
BioMed Central
23.06.2021
BioMed Central Ltd BMC |
| Subjects | |
| Online Access | Get full text |
| ISSN | 1475-2875 1475-2875 |
| DOI | 10.1186/s12936-021-03816-w |
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| Abstract | Background
Molecular and genetic studies of blood-stage
Plasmodium falciparum
parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells.
Methods
To precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth.
Results
Flow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence.
Conclusion
The combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of
P. falciparum
. This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples. |
|---|---|
| AbstractList | Abstract Background Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells. Methods To precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth. Results Flow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence. Conclusion The combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of P. falciparum. This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples. Background Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells. Methods To precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth. Results Flow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence. Conclusion The combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of P. falciparum. This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples. Keywords: Malaria, Plasmodium, Cloning, Limiting dilution Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells. To precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth. Flow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence. The combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of P. falciparum. This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples. Background Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells. Methods To precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth. Results Flow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence. Conclusion The combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of P. falciparum . This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples. Background Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells. Methods To precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth. Results Flow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence. Conclusion The combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of P. falciparum. This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples. Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells.BACKGROUNDMolecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The entire process is laborious and subject to errors due to inaccurate dilutions at the onset and failed detection of parasite growth in individual microplate wells.To precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth.METHODSTo precisely control the number of parasites dispensed into each microplate well, parasitaemia and total cell counts were determined by flow cytometry using parasite cultures stained with ethidium bromide or SYBR Green I. Microplates were seeded with 0.2 or 0.3 infected cells/well and cultivated with fresh erythrocytes. The c-SNARF fluorescent pH indicator was then used to reliably detect parasite growth.Flow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence.RESULTSFlow cytometry required less time than the traditional approach of estimating parasitaemia and cell numbers by microscopic examination. The resulting dilutions matched predictions from Poisson distribution calculations and yielded clonal lines. Addition of c-SNARF to media permitted rapid detection of parasite growth in microplate wells with high confidence.The combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of P. falciparum. This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples.CONCLUSIONThe combined use of flow cytometry for precise dilution and the c-SNARF method for detection of growth improves limiting dilution cloning of P. falciparum. This simple approach saves time, is scalable, and maximizes identification of desired parasite clones. It will facilitate DNA transfection studies and isolation of parasite clones from ex vivo blood samples. |
| ArticleNumber | 279 |
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| Author | Macedo-Silva, Tatiane Desai, Sanjay A. Wunderlich, Gerhard |
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| Cites_doi | 10.1038/nbt.2925 10.1038/nmeth.4223 10.1126/science.781840 10.1073/pnas.92.4.973 10.1016/j.jim.2011.01.015 10.1371/journal.pone.0073783 10.1126/science.7015505 10.1016/j.exppara.2008.10.008 10.1007/978-1-62703-026-7_1 10.1186/1475-2875-11-210 10.1038/nmeth1132 10.1186/1475-2875-11-124 10.1038/ncomms10727 |
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| References | W Trager (3816_CR2) 1976; 193 V Rosario (3816_CR10) 1981; 212 AG Maier (3816_CR3) 2013; 923 B Lyko (3816_CR11) 2012; 11 P Prommana (3816_CR6) 2013; 8 BT Grimberg (3816_CR13) 2011; 367 M Ghorbal (3816_CR9) 2014; 32 WHO (3816_CR1) 2019 SM Ganesan (3816_CR8) 2016; 7 CM Armstrong (3816_CR5) 2007; 4 S Izumiyama (3816_CR14) 2009; 121 S Hasenkamp (3816_CR12) 2012; 11 Y Wu (3816_CR4) 1995; 92 J Birnbaum (3816_CR7) 2017; 14 |
| References_xml | – volume: 32 start-page: 819 year: 2014 ident: 3816_CR9 publication-title: Nat Biotechnol doi: 10.1038/nbt.2925 – volume: 14 start-page: 450 year: 2017 ident: 3816_CR7 publication-title: Nat Methods doi: 10.1038/nmeth.4223 – volume: 193 start-page: 673 year: 1976 ident: 3816_CR2 publication-title: Science doi: 10.1126/science.781840 – volume: 92 start-page: 973 year: 1995 ident: 3816_CR4 publication-title: Proc Natl Acad Sci USA doi: 10.1073/pnas.92.4.973 – volume: 367 start-page: 1 year: 2011 ident: 3816_CR13 publication-title: J Immunol Methods doi: 10.1016/j.jim.2011.01.015 – volume: 8 start-page: e73783 year: 2013 ident: 3816_CR6 publication-title: PLoS One. doi: 10.1371/journal.pone.0073783 – volume: 212 start-page: 1037 year: 1981 ident: 3816_CR10 publication-title: Science doi: 10.1126/science.7015505 – volume: 121 start-page: 144 year: 2009 ident: 3816_CR14 publication-title: Exp Parasitol doi: 10.1016/j.exppara.2008.10.008 – volume: 923 start-page: 3 year: 2013 ident: 3816_CR3 publication-title: Methods Mol Biol doi: 10.1007/978-1-62703-026-7_1 – volume: 11 start-page: 210 year: 2012 ident: 3816_CR12 publication-title: Malar J doi: 10.1186/1475-2875-11-210 – volume: 4 start-page: 1007 year: 2007 ident: 3816_CR5 publication-title: Nat Methods doi: 10.1038/nmeth1132 – volume: 11 start-page: 124 year: 2012 ident: 3816_CR11 publication-title: Malar J doi: 10.1186/1475-2875-11-124 – volume: 7 start-page: 10727 year: 2016 ident: 3816_CR8 publication-title: Nat Commun doi: 10.1038/ncomms10727 – volume-title: World Malaria Report 2019 year: 2019 ident: 3816_CR1 |
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| Snippet | Background
Molecular and genetic studies of blood-stage
Plasmodium falciparum
parasites require limiting dilution cloning and prolonged cultivation in... Background Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in... Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in microplates. The... Abstract Background Molecular and genetic studies of blood-stage Plasmodium falciparum parasites require limiting dilution cloning and prolonged cultivation in... |
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| SubjectTerms | Biomedical and Life Sciences Biomedicine Blood Clones Cloning CRISPR Detection Dilution DNA Entomology Erythrocytes Ethidium bromide Experiments Flow cytometry Genes Growth Infectious Diseases Laboratories Limiting dilution Malaria Methodology Methods Microbiology Parasites Parasitology Plasmodium Plasmodium falciparum Plates Public Health Stains & staining Testing Transfection Tropical Medicine |
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| Title | Improved Plasmodium falciparum dilution cloning through efficient quantification of parasite numbers and c-SNARF detection |
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