title: Measuring Hematocrit in Mice Injected with In Vitro-Transcribed Erythropoietin mRNA.
abstract: In vitro-transcribed (IVT) mRNA encoding therapeutic protein has the potential to treat a variety of diseases by serving as template for translation in the patient. To optimize conditions for such therapy, reporter protein-encoding mRNAs are usually used. One preferred reporter is erythropoietin (EPO), which stimulates erythropoiesis and leads to an increase in hematocrit. Measurement of hematocrit is a fast and reliable method to determine the potency of the in vitro-transcribed EPO mRNA. However, frequent blood draw from mice can increase hematocrit due to blood loss. Therefore, instead of using conventional hematocrit capillary tubes, we adapted glass microcapillaries for hematocrit measurement. Daily monitoring of mice can be accomplished by drawing less than 20 μL of blood, thus avoiding blood loss-related hematocrit increase. Due to the small volume of the withdrawn blood the hematocrit remains the same for mice injected with control mRNA, whereas significant hematocrit increase is measured between day 4 and 20 postinjection for those injected with pseudouridine-modified EPO mRNA. Following hematocrit measurement the microcapillaries are snapped easily to recover plasma for further analyses, including EPO measurement by ELISA.
authors.affiliation: BioNTech RNA Pharmaceuticals GmbH, An der Goldgrube 12, Mainz, D-55131, Germany.
authors.forename: Azita Josefine
authors.initials: AJ
authors.lastname: Mahiny
authors.affiliation: BioNTech RNA Pharmaceuticals GmbH, An der Goldgrube 12, Mainz, D-55131, Germany. katalin.kariko@biontech.de.|Department of Neurosurgery, University of Pennsylvania, Philadelphia, PA, USA. katalin.kariko@biontech.de.
authors.forename: Katalin
authors.initials: K
authors.lastname: Karikó
mesh_terms: D000818:Animals; D004912:Erythrocytes / Q000378:metabolism; D004921:Erythropoietin / Q000235:genetics* / Q000378:metabolism; D006400:Hematocrit / Q000295:instrumentation*; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008807:Mice, Inbred BALB C; D012333:RNA, Messenger / Q000008:administration & dosage*
chemical_list: C554270:EPO protein, human; D012333:RNA, Messenger; D004921:Erythropoietin
keywords: Erythropoiesis; Erythropoietin (EPO); Hematocrit; In vitro-transcribed mRNA; Modified nucleosides; Protein replacement therapy; mRNA therapy