sepharose and Pancreatic-Neoplasms

We combined Michelson-interferometer-based off-axis digital holographic microscopy (DHM) with a common flow cytometry (FCM) arrangement. Utilizing object recognition procedures and holographic autofocusing during the numerical reconstruction of the acquired off-axis holograms, sharply focused quantitative phase images of suspended cells in flow were retrieved without labeling, from which biophysical cellular features of distinct cells, such as cell radius, refractive index and dry mass, can be subsequently retrieved in an automated manner. The performance of the proposed concept was first characterized by investigations on microspheres that were utilized as test standards. Then, we analyzed two types of pancreatic tumor cells with different morphology to further verify the applicability of the proposed method for quantitative live cell imaging. The retrieved biophysical datasets from cells in flow are found in good agreement with results from comparative investigations with previously developed DHM methods under static conditions, which demonstrates the effectiveness and reliability of our approach. Our results contribute to the establishment of DHM in imaging FCM and prospect to broaden the application spectrum of FCM by providing complementary quantitative imaging as well as additional biophysical cell parameters which are not accessible in current high-throughput FCM measurements.

Topics: Algorithms; Cell Line, Tumor; Female; Flow Cytometry; Holography; Humans; Image Processing, Computer-Assisted; Interferometry; Liver Neoplasms; Microscopy, Phase-Contrast; Microspheres; Middle Aged; Pancreatic Neoplasms; Refractometry; Sepharose

Isotachophoresis is an electrophoretic method of separation of charged substances. The method is characterized by a discontinuous buffer system, constant velocity of separated molecules, and the distribution of separated components in the form of narrow concentrated bands located one right after another. As a rule, isotachophoresis is not used for the separation of nucleic acids because the mobility of polynucleotides in this system does not depend on their size. However, this circumstance proved to be very useful for the quantitative isolation of heterogeneous DNA fragments from biological fluids, for gene diagnostics of cancer in particular. The proposed method of agarose gel isotachophoresis of DNA has been used for the isolation of blood DNA and its successful PCR analysis.

Topics: Buffers; DNA; DNA, Neoplasm; Electrophoresis, Agar Gel; Genetic Techniques; Humans; Neoplasms; Nucleic Acids; Pancreatic Neoplasms; Polymerase Chain Reaction; Sepharose

Topics: Animals; Biocompatible Materials; Blood Glucose; Capsules; Cell Line; Diabetes Mellitus, Experimental; Humans; Insulin Infusion Systems; Insulinoma; Islets of Langerhans Transplantation; Pancreatic Neoplasms; Rats; Rats, Inbred Lew; Sepharose; Transplantation, Heterologous; Tumor Cells, Cultured