Methods
Main content
Biobanking:
Our lab has developed biobanks of AML peripheral blood mononuclear cells (PBMCs), bone marrow cells, serum and plasma.
In addition, we have been biobanking samples from other diseases:
Sepsis (serum and plasma)
Deep Venous Thrombosis (serum and plasma)
Healthy control samples (cord blood, serum, plasma, PBMCs, and apheresis products).
We also participate in acquiring samples for different Clinical studies such as the PROLONG study (PBMC, serum, plasma, blood, feces) and a study on Cold agglutination (serum and plasma, where all handling is performed at 39 degrees before they are frozen).
Cell Culture:
We perform a lot of cell culture assays, both with primary cells, cell lines and co-cultures. To mention a few we look into drug effects, cell signaling, proliferation (3H thymidine assay), cell death, differentiation, and colony formation. We have several cell incubators, including hypoxy chambers to more closely imitate the conditions in the niches in the bone marrow. We also have a Zeiss AxioObserver FL-mik fluorescence microscope to investigate cellular localization of targets of interest.
Flow cytometry:
We have our own BD FACS Verse flow cytometer, but also use other facilities/instruments (The core facility for flow cytometry here at the institute, Section for immunology and transfusion medicine at Haukeland hospital, and the Department of Biomedical Laboratory Sciences and Chemical Engineering at the Western Norway University of Applied Sciences (HVL). We use this method in a wide variety of applications, such as cell differentiation assays, cell cycle assays, apoptosis assays, determination of cell types, quantification of cell types in blood samples, responsiveness to drugs, ROS and autophagy assays etc.
Luminex and ELISA:
We examine serum, plasma and cell culture supernatants for a wide range of components through Luminex and ELISA assays. Both assays rely upon antibody recognition of the component of interest and give you a quantified value. For example, we look into if there is a difference in the production of different cytokines in responders/non-responders to certain treatments.
Nucleic acid purification, PCR, qPCR, microarray, and sequencing:
W e look into gene mutations, and up and down regulation of genes, both in specific genes, and more non-specific screenings of cells directly from blood samples, and after mono- and co-cell cultures with and without addition of stimulators/inhibitors/cytostatics. The isolation of nucleic acids, normal and quantitative PCRs we perform ourselves, while we do the sample preparation and analyzation of the of the results from the microarray and sequencing (both gene specific and whole exome) projects.
Proteomics:
We use quantitative mass spectrometry-based proteomics to identify the proteins in a specific sample (e.g. cell lysate of primary patient samples or supernatants from cell culture) and to compare protein expression profiles between different leukemia samples. By comparing the protein expression profiles between two groups of patients (e.g. relapse/relapse-free patients), we aim to find protein biomarker candidates related to the phenotypes. We often use shotgun proteomics, which involves protein digestion and peptide separation by liquid chromatography followed by mass spectrometry analysis, and our samples are either processed with an internal standard or label-free. We have access to the mass spectrometers Orbitrap Elite and Q Exactive HF at the Proteomics Unit at the University of Bergen (PROBE). We use several bioinformatics tool to process the raw mass spectrometry data into meaningful biological information.
Seahorse:
With the Seahorse XFe96 analyzer we measure oxygen consumption rate (OCR) and Extracellular acidification rate (ECAR) of live cells in 96-well plates, and thus allowing investigation of mitochondrial respiration and glycolysis through these two key indicators. By measuring these parameters, we are provided with a system level view of metabolic function in cultured cells. The instrument is owned by the Department of Biomedical Laboratory Sciences and Chemical Engineering at the Western Norway University of Applied Sciences (HVL), with whom we collaborate.
