Malaria Methods and ProtocolsDenise L. Doolan The Plasmodium spp. parasite was identified as the causative agent of malaria in 1880, and the mosquito was identified as the vector in 1897. Despite subsequent efforts focused on the epidemiology, cell biology, immunology, molecular biology, and clinical manifestations of malaria and the Plasmodium parasite, there is still no licensed vaccine for the prevention of malaria. Physical barriers (bed nets, window screens) and chemical prevention methods (insecticides and mosquito repellents) intended to interfere with the transmission of the disease are not highly effective, and the profile of resistance of the parasite to chemoprophylactic and chemotherapeutic agents is increasing. The dawn of the new millennium has seen a resurgence of interest in the disease by government and philanthropic organizations, but we are still faced with compl- ities of the parasite, the host, and the vector, and the interactions among them. Malaria Methods and Protocols offers a comprehensive collection of protocols describing conventional and state-of-the-art techniques for the study of malaria, as well as associated theory and potential problems, written by experts in the field. The major themes reflected here include assessing the risk of infection and severity of disease, laboratory models, diagnosis and typing, molecular biology techniques, immunological techniques, cell biology techniques, and field applications. |
Contents
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25 | |
Mouse Models for ErythrocyticStage Malaria | 57 |
II Infection of Saimiri and Aotus Monkeys | 85 |
Vector Analysis | 95 |
Nested | 103 |
PCRRFLP Analysis | 117 |
Microsatellite Analysis in Plasmodium falciparum | 131 |
Peptide Vaccination | 335 |
DNA Vaccination | 347 |
Assessing AntigenSpecific CD8+ and CD4+TCell Responses in Mice | 361 |
Assessing CD4+ Helper TLymphocyte Responses by Lymphoproliferation | 369 |
Limiting Dilution Analysis of AntigenSpecific CD4+ TCell Responses | 385 |
Malaria BloodStage ParasiteSpecific CD4+T Cells After | 401 |
Using Flow Cytometry | 409 |
Aftab A Ansari and Ann E Mayne | 423 |
Quantitation of LiverStage Parasites | 137 |
Extraction and Purification of Plasmodium Total | 151 |
Extraction and Purification of Plasmodium Parasite | 159 |
SDSPAGE and Western Blotting of Plasmodium falciparum Proteins | 177 |
Nested PCR Analysis of Plasmodium Parasites | 189 |
RFLP Analysis | 205 |
Analysis of Gene Expression by RTPCR | 213 |
In Situ Detection of RNA in Blood and MosquitoStage Malaria Parasites | 225 |
Daniel J Carucci Paul Horrocks and Malcolm J Gardner 23 Construction of Genomic Libraries from the DNA of Plasmodium Species | 235 |
Cummings Dharmendar Rathore | 241 |
NucleaseTreated Genomic | 253 |
David A Fidock Dharmendar Rathore and Thomas F McCutchan | 265 |
Production of StageSpecific Plasmodium falciparum cDNA Libraries | 277 |
Construction and Screening of YAC Libraries | 291 |
Episomal Transformation of Plasmodium berghei | 305 |
Vandana Thathy and Robert Ménard | 317 |
and José Alejandro López 39 Assessing AntigenSpecific CD8+ CTL Responses in Humans | 437 |
Doolan | 445 |
Human Antibody Subclass ELISA | 457 |
II Analysis | 469 |
In Vitro Culture of Plasmodium Parasites | 477 |
J David Haynes and J Kathleen Moch 45 Hepatic Portal Branch Inoculation | 489 |
John B Sacci Jr | 499 |
Hepatocyte Perfusion Isolation and Culture | 503 |
Inhibition of LiverStage Development Assay | 517 |
Erythrocytic Malaria Growth or Invasion Inhibition Assays with Emphasis | 535 |
Analysis of CSABinding Parasites and Antiadhesion Antibodies | 555 |
Analysis of the Adhesive Properties of Plasmodium falciparumInfected | 561 |
Preparation of Adhesive Targets for FlowBased Cytoadhesion Assays | 571 |
Triton X114 Phase Partitioning for Antigen Characterization | 581 |
Field Trials | 607 |
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Common terms and phrases
acid agarose aliquot alleles amplification analysis antibodies antigen approx assay buffer cDNA Centrifuge chabaudi clones concentration containing culture medium cycle cytokines detection diluted EDTA epitopes erythrocytes ethanol ethidium bromide filter flasks flow cytometry fragments gene genomic DNA hepatocytes human hybridization immune Immunol Incubate infection inhibition labeled liver Malaria Methods malaria parasites marker membrane merozoite Methods and Protocols mg/mL mice microslides Molecular monkeys mosquitoes mouse Nest Note parasite parasitemia Parasitol PCR product pellet peptide pipet plasmid Plasmodium berghei Plasmodium falciparum Plasmodium yoelii plate polymerase Prepare primers procedure protein purification reaction reagents recombinant remove Resuspend room temperature RPMI sample schizonts sequences serum Sigma slides sodium solution specific sporozoites stages staining sterile Store at 4°C Subheading supernatant syringe T-cell target template tion transfer Tris-HCl tube vaccine vector vitro volume wash yoelii µg/mL