Skip to main content

Advertisement

SpringerLink
Log in

Production of monoclonal antibodies for Plasmodium vivax lactate dehydrogenase and patient sera screening using sandwich ELISA

  • Original Paper
  • Published:
Parasitology Research Aims and scope Submit manuscript

Abstract

Malaria is a worldwide infectious disease. There are many diagnostic kits to detect malaria infection. However, the sensitivity of these diagnostic kits remains a problem. To develop a diagnostic kit for malaria that has high sensitivity, it is necessary to produce monoclonal antibodies (McAbs) with high affinity. The present study was undertaken to produce hybridoma cells that can be used to generate McAbs with high affinity and specificity against Plasmodium vivax lactate dehydrogenase (pvLDH). In this study, BALB/c mice were immunized with purified recombinant polypeptides that encode pvLDH. McAbs against pvLDH were produced according to the protocol of hybridoma technique using myeloma cells (SP2/0 cell lines). The McAbs were characterized by isotyping and by Western blot analysis. Two McAbs (D2H and D7E) against pvLDH antigen were obtained. The isotypes of D2H and D7E were IgG2b. They recognize 33 kDa proteins that were defined as pvLDH by Western blot analysis. In the affinity test, D2H and D7E showed positively optical density value until each McAbs were serially diluted at concentrations of 0.156 and 0.078 μg/ml, respectively. To evaluate sensitivity and specificity against clinical specimens of P. vivax, purified McAbs were tested with alkaline phosphatase-conjugated monoclonal antibodies and blood samples (n = 180) of P. vivax patients using the sandwich enzyme-linked immunosorbent assay, showing the 98 % sensitivity. We suggest that McAbs produced in this study may be used for developing efficient and rapid diagnostic kits.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Campbell GH, Miller LH, Hudson D, Franco EL, Andrysiak PM (1984) Monoclonal antibody characterization of Plasmodium falciparum antigens. Am J Trop Med Hyg 33:1051–1054

    PubMed  CAS  Google Scholar 

  • Cho D, Kim KH, Park SC, Kim YK, Lee KN, Lim CS (2001) Evaluation of rapid immunocapture assays for diagnosis of Plasmodium vivax in Korea. Parasitol Res 87:445–448

    Article  PubMed  CAS  Google Scholar 

  • Coleman RE, Maneechai N, Ponlawat A, Kumpitak C, Rachapaew N, Miller RS, Sattabongkot J (2002) Short report: failure of the OptiMAL rapid malaria test as a tool for the detection of asymptomatic malaria in an area of Thailand endemic for Plasmodium falciparum and P. vivax. Am J Trop Med Hyg 67:563–565

    PubMed  Google Scholar 

  • Congpuong K, Na-Bangchang K, Thimasarn K, Tasanor U, Wernsdorfer WH (2002) Sensitivity of Plasmodium vivax to chloroquine in Sa Kaeo Province, Thailand. Acta Trop 83:117–121

    Article  PubMed  CAS  Google Scholar 

  • Cooke AH, Chiodini PL, Doherty T, Moody AH, Ries J, Pinder M (1999) Comparison of a parasite lactate dehydrogenase-based immunochromatographic antigen detection assay (OptiMAL) with microscopy for the detection of malaria parasites in human blood samples. Am J Trop Med Hyg 60:173–176

    PubMed  CAS  Google Scholar 

  • Doolan DL, Southwood S, Freilich DA, Sidney J, Graber NL, Shatney L, Bebris L, Florens L, Dobano C, Witney AA, Appella E, Hoffman SL, Yates JR 3rd, Carucci DJ, Sette A (2003) Identification of Plasmodium falciparum antigens by antigenic analysis of genomic and proteomic data. Proc Natl Acad Sci U S A 100:9952–9957

    Article  PubMed  CAS  Google Scholar 

  • Ferro BE, Gonzalez IJ, Carvajal F, Palma GI, Saravia NG (2002) Performance of OptiMAL in the diagnosis of Plasmodium vivax and Plasmodium falciparum infections in a malaria referral center in Colombia. Mem Inst Oswaldo Cruz 97:731–735

    Article  PubMed  Google Scholar 

  • Florens L, Washburn MP, Raine JD, Anthony RM, Grainger M, Haynes JD, Moch JK, Muster N, Sacci JB, Tabb DL, Witney AA, Wolters D, Wu Y, Gardner MJ, Holder AA, Sinden RE, Yates JR, Carucci DJ (2002) A proteomic view of the Plasmodium falciparum life cycle. Nature 419:520–526

    Article  PubMed  CAS  Google Scholar 

  • Forney JR, Magill AJ, Wongsrichanalai C, Sirichaisinthop J, Bautista CT, Heppner DG, Miller RS, Ockenhouse CF, Gubanov A, Shafer R, DeWitt CC, Quino-Ascurra HA, Kester KE, Kain KC, Walsh DS, Ballou WR, Gasser RA Jr (2001) Malaria rapid diagnostic devices: performance characteristics of the ParaSight F device determined in a multisite field study. J Clin Microbiol 39:2884–2890

    Article  PubMed  CAS  Google Scholar 

  • Galfrè G, Milstein C (1981) Preparation of monoclonal antibodies: strategies and procedures. Methods Enzymol 73:3–46

    Article  PubMed  Google Scholar 

  • Gaye O, Diouf M, Diallo S (1999) A comparison of thick smears, QBC malaria, PCR and PATH falciparum malaria test trip in Plasmodium falciparum diagnosis. Parasite 6:273–275

    PubMed  CAS  Google Scholar 

  • Kim KH, Jang JW, Woo MK, Oh JS, Han ET, Lee WJ, An SS, Lim CS (2011) Evaluation of four rapid diagnostic tests for the diagnosis of Plasmodium vivax in Korea. Trop Med Int Health 16:1427–1431

    Article  PubMed  Google Scholar 

  • Köhler G, Milstein C (1975) Continuous cultures of fused cells secreting antibody of predefined specificity. Nature 256:495–497

    Article  PubMed  Google Scholar 

  • Le Roch KG, Zhou Y, Blair PL, Grainger M, Moch JK, Haynes JD, De La Vega P, Holder AA, Batalov S, Carucci DJ, Winzeler EA (2003) Discovery of gene function by expression profiling of the malaria parasite life cycle. Science 301:1503–1508

    Article  PubMed  Google Scholar 

  • Lee SW, Jeon K, Jeon BR, Park I (2008) Rapid diagnosis of vivax malaria by SD Bioline malaria antigen test when thrombocytopenia is present. J Clin Microbiol 46:939–942

    Article  PubMed  Google Scholar 

  • Lim KJ, Park JW, Sohn MJ, Lee S, Oh JH, Kim HC, Bahk YY, Kim YS (2002) A direct sandwich ELISA to detect antibodies against the C-terminal region of merozoite surface protein 1 could be a useful diagnostic method to identify Plasmodium vivax exposed persons. Parasitol Res 88:855–860

    Article  PubMed  Google Scholar 

  • Macreadie I, Ginsburg H, Sirawaraporn W, Tilley L (2000) Antimalarial drug development and new targets. Parasitol Today 16:438–444

    Article  PubMed  CAS  Google Scholar 

  • Makler MT, Hinrichs DJ (1993) Measurement of the lactate dehydrogenase activity of Plasmodium falciparum as an assessment of parasitemia. Am J Trop Med Hyg 48:205–210

    PubMed  CAS  Google Scholar 

  • Makler MT, Ries JM, Williams JA, Bancroft JE, Piper RC, Gibbins BL, Hinrichs DJ (1993) Parasite lactate dehydrogenase as an assay for Plasmodium falciparum drug sensitivity. Am J Trop Med Hyg 48:739–741

    PubMed  CAS  Google Scholar 

  • Makler MT, Palmer CJ, Ager AL (1998) A review of practical techniques for the diagnosis of malaria. Ann Trop Med Parasitol 92:419–433

    Article  PubMed  CAS  Google Scholar 

  • Mason DP, Kawamoto F, Lin K, Laoboonchai A, Wongsrichanalai C (2002) A comparison of two rapid field immunochromatographic tests to expert microscopy in the diagnosis of malaria. Acta Trop 82:51–59

    Article  PubMed  Google Scholar 

  • Moody A (2002) Rapid diagnostic tests for malaria parasites. Clin Microbiol Rev 15:66–78

    Article  PubMed  CAS  Google Scholar 

  • Moody A, Hunt-Cooke A, Gabbett E, Chiodini P (2000) Performance of the OptiMAL malaria antigen capture dipstick for malaria diagnosis and treatment monitoring at the Hospital for Tropical Diseases, London. Br J Haematol 109:891–894

    Article  PubMed  CAS  Google Scholar 

  • Murray CK, Bell D, Gasser RA Jr, Wongsrichanalai C (2003) Rapid diagnostic testing for malaria. Trop Med Int Health 8:876–883

    Article  PubMed  Google Scholar 

  • Mya MM, Roy A, Saxena RK, Roy KB (2002) Isolation, part characterization, immunogenicity, and specificity study of Plasmodium falciparum culture supernatant. Jpn J Infect Dis 55:150–156

    PubMed  Google Scholar 

  • Park JW, Yoo SB, Oh JH, Yeom JS, Lee YH, Bahk YY, Kim YS, Lim KJ (2008) Diagnosis of vivax malaria using an IgM capture ELISA is a sensitive method, even for low levels of parasitemia. Parasitol Res 103:625–631

    Article  PubMed  Google Scholar 

  • Pinto MJ, Pereira NF, Rodrigues S, Kharangate NV, Verenkar MP (1999) Rapid diagnosis of falciparum malaria by detection of Plasmodium falciparum HRP-2 antigen. J Assoc Physicians India 47:1076–1078

    PubMed  CAS  Google Scholar 

  • Piper RC, Vanderjagt DL, Holbrook JJ, Makler M (1996) Malaria lactate dehydrogenase: target for diagnosis and drug development. Ann Trop Med Parasitol 90:433

    Google Scholar 

  • Richter J, Göbels K, Müller-Stöver I, Hoppenheit B, Häussinger D (2004) Co-reactivity of plasmodial histidine-rich protein 2 and aldolase on a combined immuno-chromographic-malaria dipstick (ICT) as a potential semi-quantitative marker of high Plasmodium falciparum parasitaemia. Parasitol Res 94:384–385

    Article  PubMed  Google Scholar 

  • Rogerson SJ, Carter R (2008) Severe vivax malaria: newly recognised or rediscovered. PLoS Med 5:e136

    Article  PubMed  Google Scholar 

  • Rubio JM, Benito A, Berzosa PJ, Roche J, Puente S, Subirats M, López-Vélez R, García L, Alvar J (1999) Usefulness of seminested multiplex PCR in surveillance of imported malaria in Spain. J Clin Microbiol 37:3260–3264

    PubMed  CAS  Google Scholar 

  • Vander Jagt DL, Hunsaker LA, Heidrich JE (1981) Partial purification and characterization of lactate dehdrogenase from Plasmodium falciparum. Mol Biochem Parasitol 4:255–264

    Article  PubMed  CAS  Google Scholar 

  • Winter VJ, Cameron A, Tranter R, Sessions RB, Brady RL (2003) Crystal structure of Plasmodium berghei lactate dehydrogenase indicates the unique structural differences of these enzymes are shared across the Plasmodium genus. Mol Biochem Parasitol 131:1–10

    Article  PubMed  CAS  Google Scholar 

  • Wongsrichanalai C (2001) Rapid diagnostic techniques for malaria control. Trends Parasitol 17:307–309

    Article  PubMed  CAS  Google Scholar 

  • Wongsrichanalai C, Gasser RA Jr (2002) Current status of malaria rapid diagnostic devices: an update. Trends Parasitol 18:107–108

    Article  PubMed  Google Scholar 

  • Wongsrichanalai C, Barcus MJ, Muth S, Sutamihardja A, Wernsdorfer WH (2007) A review of malaria diagnostic tools: microscopy and rapid diagnostic test (RDT). Am J Trop Med Hyg 77(6 Suppl):119–127

    PubMed  Google Scholar 

  • World Health Organization (2010a) WHO guidelines for the treatment of malaria, 2nd edn. WHO, Geneva

    Google Scholar 

  • World Health Organization (2010b) Malaria rapid diagnostic test performance. Results of WHO product testing of malaria RDTs: round 2, 2009. WHO, Geneva

    Google Scholar 

Download references

Acknowledgments

This research was supported by a Research fund (2011-E54009-00) of the Korea Centers for Disease Control and Prevention, and by the Public Welfare & Safety Research Program through the National Research Foundation of Korea funded by the Ministry of Education, Science and Technology (2011-0020967).

Author information

Authors and Affiliations

  1. Department of Microbiology, and Molecular Science and Technology, Ajou University School of Medicine, Suwon, 443-721, Republic of Korea

    Jong-Hyun Kim, Jinyoung Lee, Hae-Jin Sohn & Ho-Joon Shin

  2. Department of Infection Biology, Wonkwang University College of Medicine, Iksan, 570-749, Republic of Korea

    Hyun-Ok Song & Hyun Park

  3. Division of Malaria and Parasitic Diseases, Korea National Institute of Health, Osong, 363-951, Republic of Korea

    Jung-Yeon Kim & Won-Ja Lee

Authors
  1. Jong-Hyun Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jinyoung Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hae-Jin Sohn
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hyun-Ok Song
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jung-Yeon Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Won-Ja Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Hyun Park
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Ho-Joon Shin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Hyun Park or Ho-Joon Shin.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kim, JH., Lee, J., Sohn, HJ. et al. Production of monoclonal antibodies for Plasmodium vivax lactate dehydrogenase and patient sera screening using sandwich ELISA. Parasitol Res 111, 1645–1650 (2012). https://doi.org/10.1007/s00436-012-3003-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00436-012-3003-x

Keywords

Search

Navigation