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Detection of Metastatic Tumor Cells in Bone Marrow
Published in Adrian P. Gee, BONE MARROW PROCESSING and PURGING, 2020
Another method for detection of minimal marrow disease is the use of the clonogenic assay. This technique, first reported by Hamburger and Salmon,30 was an attempt to define poor prognostic malignancies by analyzing their potential for colony growth in vitro. Several investigators have used a modification of this method to grow neoplastic colonies from bone marrow.17,31–34 This has been proven to be superior to routine histologic analysis. However, the number of tumor cells capable of producing colonies in this assay (plating efficiency) is low, even if highly malignant cells are used (e.g., neoplastic cell lines).35,36 Consequently, the sensitivity of this method is low, and studies have shown that immunologic detection techniques are superior. Despite this drawback, the clonogenic assay is useful for malignancies where monoclonal antibodies that can differentiate tumor cells from normal hematopoietic cells are unavailable (e.g., in Hodgkin’s and non-Hodgkin’s lymphoma).32
Assay of Antibiotics in Mammalian Cell Culture
Published in Adorjan Aszalos, Modern Analysis of Antibiotics, 2020
Recently, the human tumor clonogenic assay emerged as the tissue culture system offering the most promise for the discovery of substances active against human tumors. Its application to large-scale screening is still in the developmental stage. However, its usefulness as a predictive test for the chemotherapeutic treatment of cancer patients has met with some success in a limited number of tumor types. Although it is not expected that this assay will replace well-standardized methods already established for potency and other routine cytotoxicity evaluations, or most mechanisms of action studies, it may have a place in defining activities heretofore not seen in cytotoxicity studies using continuous cell lines.
Prolactin Receptors in Normal Tissues and in Animal Models for Breast Cancer
Published in Nagasawa Hiroshi, Prolactin and Lesions in Breast, Uterus, and Prostate, 2020
Paul A. Kelly, David Gould, Hiroaki Okamura, Jean Dijane
The clonogenic assay is a method in which biopsy specimens are dissociated to single cells and then cultured in soft agar. This procedure not only measures the growth stimulating activity of a hormone treatment, but also identifies conditions which stimulate colony formation from single cells, a fundamental characteristic of cancer cells. In the NMU rat tumor, both ovine PRL and ovine GH stimulate colony formation.164 In human tumors, PRL alone stimulated malignant tumor growth in nine of ten estrogen receptor-positive tumors and three of seven estrogen receptor-negative tumors.165
3D high resolution clonogenic survival measurement of xrs-5 cells in low-dose region of carbon ion plans
Published in International Journal of Radiation Biology, 2023
Dea Kartini, Olga Sokol, Chutima Talabnin, Chinorat Kobdaj, Marco Durante, Michael Krämer, Martina Fuss
For recovering cells from matrigel, the medium layer was discarded and 22 µL of dispase (Corning) was added and incubated for 1 h to dissolve the matrigel matrix. The dissolved gel was collected into microtubes which were filled up with medium until the total volume of suspension reached 1 mL and then centrifuged for 5 min (2500 rpm, 4µL of trypsin was mixed with the cell pellet and incubated for 3 min to separate cell clumps into single cells. Afterwards, 900 µL of medium was added to deactivate the trypsin. Cells were counted using the Beckman Coulter counter using a profile with 1:50 dilution and 7 − 18 µm particle size. The average number of cells obtained was 75,048 cells per well with an uncertainty of ±11.5%. We find the cell number obtained is sufficient for performing the clonogenic assay.
Radiation impacts on toxicity of cobalt–chromium (CoCr) implant debris
Published in Nanotoxicology, 2023
Kevin L. Trout, Sanghamitra Majumdar, Anil K. Patri, Tariq Fahmi
RTCA is a label-free method for observing changes in cell adhesion, morphology, proliferation, and death that are reflected by cell index readings. This technique allows for continuous monitoring of cells, while other assays in this study rely on endpoint measurements. RTCA requires cells to be seeded at the same initial density to calibrate cell index values and enable comparisons among treatment groups. Curves of untreated cells increase until cells reach confluence, then they begin to decline. Higher cell index values were observed in HMEC-1, likely because these endothelial cells can form a tighter monolayer than SW982. Readings were recorded for 6 d; data beyond this point were less reliable due to over-growth of untreated cells and reduced survival of treated cells. This limitation is avoided in the clonogenic assay by adjusting cell plating numbers to allow for an extended timeline. Nonetheless, RTCA data supported the conclusions from both the short timeline viability assays and extended timeline clonogenic assays (Figure 9). CoCr and radiation resulted in lower normalized cell indices over time, indicating lower cell survival.
Polydatin-Induced Direct and Bystander Effects in A549 Lung Cancer Cell Line
Published in Nutrition and Cancer, 2022
To check the anticancer activity, cells were treated with 0–1 mM concentration of polydatin for 72 h. The normal morphology of untreated A549 cells is spindle shaped but drug treated groups (0.2–0.6 mM) showed flat and enlarged cells (Figure 1a) with increased nuclear size (Figure 1b). At higher concentration (>0.6 mM), cells became thin and elongated with lots of projections (Figure 1a). Number of floating/dead cells also increased corresponding to increased drug concentration (0.8–1 mM). Clonogenic assay further showed reduced number and size of colonies. The number of colonies reduced to about 50% of control at 0.2 mM concentration. At higher concentration of polydatin, the colony forming ability gradually decreased and no colony was observed at 0.8 mM (Figure 1c). Cell density also reduced significantly after 72 h of polydatin treatment (Figure 1d). This indicated that polydatin induces growth inhibition in a dose dependent manner. The expression of Ki67 protein also decreased significantly with increase in drug concentration (0.8–1 mM) (Figure 1e). These results show that polydatin inhibits cell growth at lower concentration and induces cell death at higher concentration.