SYBR® Green I-Based Fluorescence Assay to Assess Cell Viability of Malaria Parasites for Routine Use in Compound Screening

Leidenberger, Maria; Voigtländer, Cornelia; Simon, Nina; Kappes, Barbara

doi:10.1007/978-1-4939-6960-9_9

Cited by 14 publications

(11 citation statements)

References 21 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Relative parasite proliferation was determined either using a SYBR Green I-based fluorescence assay ( Leidenberger et al, 2017 ) or by flow cytometry. For flow cytometry, cultures were fixed in PBS containing 4% formaldehyde and 0.0075% glutaraldehyde and stained with 20 µM Hoechst 33,342 (Invitrogen) and 1 μg/ml thiazole orange (Sigma Aldrich) for 30 min.…”

Section: Methodsmentioning

confidence: 99%

The Putative Bromodomain Protein PfBDP7 of the Human Malaria Parasite Plasmodium Falciparum Cooperates With PfBDP1 in the Silencing of Variant Surface Antigen Expression

Quinn

Jeninga

Limm

et al. 2022

Front. Cell Dev. Biol.

View full text Add to dashboard Cite

Epigenetic regulation is a critical mechanism in controlling virulence, differentiation, and survival of the human malaria parasite Plasmodium (P.) falciparum. Bromodomain proteins contribute to this process by binding to acetylated lysine residues of histones and thereby targeting the gene regulatory machinery to gene promoters. A protein complex containing the P. falciparum bromodomain proteins (PfBDP) 1 and PfBDP2 (BDP1/BDP2 core complex) was previously shown to play an essential role for the correct transcription of invasion related genes. Here, we performed a functional characterization of a third component of this complex, which we dubbed PfBDP7, because structural modelling predicted a typical bromodomain fold. We confirmed that PfBDP7 is a nuclear protein that interacts with PfBDP1 at invasion gene promoters in mature schizont stage parasites and contributes to their transcription. Although partial depletion of PfBDP7 showed no significant effect on parasite viability, conditional knock down of either PfBDP7 or PfBDP1 resulted in the de-repression of variant surface antigens (VSA), which are important pathogenicity factors. This de-repression was evident both on mRNA and protein level. To understand the underlying mechanism, we mapped the genome wide binding sites of PfBDP7 by ChIPseq and showed that in early schizonts, PfBDP7 and PfBDP1 are commonly enriched in heterochromatic regions across the gene body of all VSA families, including genes coding for PfEMP1, RIFIN, STEVOR, and PfMC-2TM. This suggests that PfBDP7 and PfBDP1 contribute to the silencing of VSAs by associating with heterochromatin. In conclusion, we identified PfBDP7 as a chromatin binding protein that is a constitutive part of the P. falciparum BDP1/BDP2 core complex and established PfBDP1 and PfBDP7 as novel players in the silencing of heterochromatin regulated virulence gene families of the malaria parasite P. falciparum.

show abstract

Section: Methodsmentioning

confidence: 99%

The Putative Bromodomain Protein PfBDP7 of the Human Malaria Parasite Plasmodium Falciparum Cooperates With PfBDP1 in the Silencing of Variant Surface Antigen Expression

Quinn

Jeninga

Limm

et al. 2022

Front. Cell Dev. Biol.

View full text Add to dashboard Cite

show abstract

“…The screening of the antiplasmodial activities of the five herbal formulations was performed using the SYBR green I-based fluorescence assay as previously described. 14,15 The infected Red blood cell were incubated with herbal formulation and artesunate under culture conditions using 1 % parasitaemia and 2 % hematocrit. The negative controls were not treated with any drug.…”

Section: Assessment Of Antiplasmodial Activitymentioning

confidence: 99%

“…The MTT 3-(4, 5-dimethylthiazol-2-yl)-5-diphenyltetrazolium bromide-based assay as described in earlier studies with slight modification was used for this study. 14,15 Briefly, for erythrocyte survival assay, a 100 μL of washed Red blood cell at 2% hematocrit was added 100 μL volumes of serially diluted herbal formulations (For each herbal formulation, five duplicate dilutions were prepared with concentrations ranging from 6.25 μg/mL to 100 μg/mL. The plates were incubated at 37 0 C for 72 hours in a modular chamber under low oxygen and carbon dioxide.…”

Section: Screening Of Herbal Formulations For Red Blood Cell Cytotoxi...mentioning

confidence: 99%

Screening of Five Herbal Formulations Sold in South-East Nigeria for their Phytochemical Properties, In Vitro Antioxidant, Antiplasmodial and Cytotoxic Activities

2022

TJNPR

View full text Add to dashboard Cite

such as phenols, flavonoids and tannins which have gained more recognition for their role in the prevention of human diseases. 8 The evaluated herbal formulations are claimed by the manufacturers to have so much medicinal importance and can be used to treat several ailments, including malaria, typhoid, and serve as antioxidants, antibacterial, and antifungal remedies in traditional medicine practice in Nigeria. However, there has not been any scientific validation to back up these claims. The present study assessed the qualitative phytochemical properties, in vitro antioxidant, antiplasmodial and cytotoxicity effect of these herbal formulations sold in Nigeria. Materials and Methods Parasites strainThe (DD2) chloroquine resistant strain of the plasmodium parasite was acquired from the

show abstract

“…These methods include DNA-binding fluorescent dyes [242], the parasite lactate dehydrogenase assay methods [243] or the use of transgenic parasites expressing luciferase-reporter cassettes [244][245][246][247]. The SYBR ® green fluorescence-based screening with blood stage P. falciparum cultures has been the hallmark of antimalarial drug discovery for more than a decade [248]. SYBR ® green-based assays have been further optimized for antimalarial screening under low levels of parasitemia against clinical field isolates of P. falciparum for surveys of drug resistance [249,250] in infections with mixed strains and for the evaluation of antimalarial drug combinations [251].…”

Section: Phenotypic Vs Target-based Screening In Malariamentioning

confidence: 99%

Screening Marine Natural Products for New Drug Leads against Trypanosomatids and Malaria

et al. 2020

View full text Add to dashboard Cite

Neglected Tropical Diseases (NTD) represent a serious threat to humans, especially for those living in poor or developing countries. Almost one-sixth of the world population is at risk of suffering from these diseases and many thousands die because of NTDs, to which we should add the sanitary, labor and social issues that hinder the economic development of these countries. Protozoan-borne diseases are responsible for more than one million deaths every year. Visceral leishmaniasis, Chagas disease or sleeping sickness are among the most lethal NTDs. Despite not being considered an NTD by the World Health Organization (WHO), malaria must be added to this sinister group. Malaria, caused by the apicomplexan parasite Plasmodium falciparum, is responsible for thousands of deaths each year. The treatment of this disease has been losing effectiveness year after year. Many of the medicines currently in use are obsolete due to their gradual loss of efficacy, their intrinsic toxicity and the emergence of drug resistance or a lack of adherence to treatment. Therefore, there is an urgent and global need for new drugs. Despite this, the scant interest shown by most of the stakeholders involved in the pharmaceutical industry makes our present therapeutic arsenal scarce, and until recently, the search for new drugs has not been seriously addressed. The sources of new drugs for these and other pathologies include natural products, synthetic molecules or repurposing drugs. The most frequent sources of natural products are microorganisms, e.g., bacteria, fungi, yeasts, algae and plants, which are able to synthesize many drugs that are currently in use (e.g. antimicrobials, antitumor, immunosuppressants, etc.). The marine environment is another well-established source of bioactive natural products, with recent applications against parasites, bacteria and other pathogens which affect humans and animals. Drug discovery techniques have rapidly advanced since the beginning of the millennium. The combination of novel techniques that include the genetic modification of pathogens, bioimaging and robotics has given rise to the standardization of High-Performance Screening platforms in the discovery of drugs. These advancements have accelerated the discovery of new chemical entities with antiparasitic effects. This review presents critical updates regarding the use of High-Throughput Screening (HTS) in the discovery of drugs for NTDs transmitted by protozoa, including malaria, and its application in the discovery of new drugs of marine origin.

show abstract

SYBR® Green I-Based Fluorescence Assay to Assess Cell Viability of Malaria Parasites for Routine Use in Compound Screening

Cited by 14 publications

References 21 publications

The Putative Bromodomain Protein PfBDP7 of the Human Malaria Parasite Plasmodium Falciparum Cooperates With PfBDP1 in the Silencing of Variant Surface Antigen Expression

The Putative Bromodomain Protein PfBDP7 of the Human Malaria Parasite Plasmodium Falciparum Cooperates With PfBDP1 in the Silencing of Variant Surface Antigen Expression

Screening of Five Herbal Formulations Sold in South-East Nigeria for their Phytochemical Properties, In Vitro Antioxidant, Antiplasmodial and Cytotoxic Activities

Screening Marine Natural Products for New Drug Leads against Trypanosomatids and Malaria

Contact Info

Product

Resources

About