Frequently Asked Questions (FAQ) for Inverted Microscope
1. How could I fully know about Cell therapy from The Point of Standardization, Scale, and Industrialization?What is cell therapy?Cell therapy refers to the transplantation or input of normal or bioengineered human cells into a patient's body and newly-imported cells can replace damaged cells or involve a stronger immune killing function, so as to achieve the purpose of treating diseases. Cell therapy has shown higher application value in the treatment of cancer, hematological diseases, cardiovascular diseases, diabetes, Alzheimer's disease etc. In general, cell therapy includes tumor cell immunotherapy and stem cell therapy. There are two cell sources for cell therapy, one from the patient itself and the other from the allogeneic tissue.The Defects of Cell TherapyThe cell is the most basic unit that contributes to a living organism, however, it does not mean that everyone shares the same cells. On the contrary, there is a huge difference in each individual which can be compared to human-to-human differences, that is, two identical people never exist. The huge difference between cells and cell preparations is the biggest drawback of cell therapy. In this post, we will discuss several issues that need attention in the current stage of cell therapy.Difficulties in the Standardization of Cell TherapyCancer cell immunotherapy cannot be standardized from the stage of raw material acquisition. The cell treatment materiasl for each paitient are their own blood leukocytes. The condition and physical condition of each patient are different, and the collected white blood cell growth quantity and kill activity are not uniform and cannot be standardized. As it is impossible to standardize raw materials, preparation processes, and product specifications, it cannot be standardized, industrialized, and scaled up. Each tumor cell immunotherapy laboratory meets the GMP level with the hardware environment, and it can be more like a cell preparation workshop. Researchers ranged in number from a few to a dozen and could not really meet the standards of division of labor of industrialized pharmaceutical companies. Taking stem cell therapy that using umbilical cord mesenchymal stem cells as an example, which raw material is an umbilical cord, and one umbilical cord-produced cell can be utilized by many paitients. The standardization path is more advanced than the immunotherapy of tumor cells, and the raw materials can be standardized to some extent.Difficulties in The Scale of Cell Therapy IndustryAt present, the production mode of the cell therapy industry mainly depends on technicians. In the 10,000-grade clean laboratory, the cells are operated in a class 100 clean bench, cultured in a carbon dioxide incubator, centrifuged in a centrifuge, observed through an inverted microscope, and the drug reagents are stored in a medicine refrigerator. All of these devices are operated by independent biological laboratories of the individual and being linked together through the operations of scientists. This type of production model is small in scale and similar to workshop-type production. Although there are some large scales, the essence is a collection of many small workshops. Due to the small scale, the instruments used are laboratory instruments and many of the reagents used are scientific reagents, which will lead to the issue of low efficiency but high cost.Autologous or Allogeneic cellsThere are two kinds of cell sources for cell therapy, one from the patients and the other from the allogeneic tissue. Autologous cell therapy can not be standardized from the raw material acquisition stage, and its cells are only applied to the patient itself, the essence is essentially medical technology. The prevalence of autologous cell therapy as a medical technology is mainly due to the scale of the predicament. Allogeneic cell therapy, the cells derived from allogeneic. Taking tumor cell immunotherapy as an example, the cell source may be from cord blood, and the larger-scale cell source may be a filter plate for leukocyte filtration at the blood bank. Taking umbilical cord mesenchymal stem cells as an example, the cell source is the umbilical cord, and one umbilical cord-producing cell can be used by more than one person. If scale can be cultivated, although the quality standards cannot be quantified well, the scaled products themselves have a certain degree of standardized properties.The cell industry, as an industry, is not the path to the advancement of cell-based therapeutics. If the advanced technology cannot be mass-produced on a large scale, it can only stay in the laboratory and become the object of research for scientists, never have a chance to become a drug into the majority of patients. For allogeneic cell therapy that using allogeneic cells as raw materials, the standardized properties of the scaled products can be realized if large-scale cultures are prepared, then scale and standardization can promote each other. The current progress in standardization of cells is not easy, but the progress in scale should be relatively easy to achieve. Natural cytokine supernatants with more standardized and standardized propertiesCytokines are a class of small molecule proteins with broad biological activity synthesized and secreted by immune cells (such as monocytes, macrophages, T cells, B cells, NK cells, etc.) and certain non-immune cells (endothelial cells, epidermal cells, fibroblasts, etc.) Immune responses are regulated by binding to the respective receptors to regulate cell growth, differentiation and effects. Cytokines (CK) are low-molecular-weight soluble proteins that are produced by various types of cells induced by immunogens, mitogens, or other stimulants. They have the ability to regulate innate immunity and adaptive immunity , hematopoiesis, cell growth, and damage tissue repair and other functions.Cytokines can be divided into interleukins, interferons, tumor necrosis factor superfamily, colony stimulating factors, chemokines, growth factors etc. Cytokines form a very complex cytokine regulatory network in the body and participate in many important physiological functions of the human body. Where stem cells and immune cells cannot reach the body, cytokines can easily reach target tissue sites because of their small size.In recent years, recombinant gene cytokines have made remarkable achievements in clinical applications as a novel biological response modifier. A large part of the effects of stem cell therapy and immune cell therapy arises from the action of cytokines secreted in the body. The stem cells and immune cells in the body are introduced back into the body to secrete a variety of natural structural cytokines. Although the amount of these cytokines is relatively small, they are synergistic and act directly on the cytokine network in the body because of their high natural structure activity, lack of antigenicity but diversity. Because of the standardization, standardization, industrialization, and scale of natural compound cytokines, it is more cost-effective than cell therapy, allowing more patients in need to enjoy cell-like therapeutic effects.Although natural complex cytokines can largely replace cell therapy, but there are still conditions that require the presence of cells to exert a therapeutic effect. We hope that cell therapy can break the current situation, become high efficiency and low cost with large scale, more standardization, and then be applied to more disease treatments. How could I fully know about Cell therapy from The Point of Standardization, Scale, and Industrialization?------2. What are some easy ways to store freshwater samples from ponds, preserving the life forms (microorganisms like plankton, algae)? Would freezing work?It depends on what you want to do with the samples, but freezing tends to cause ice crystals form in the tissues, rupturing the cells. I donu2019t know if there is an easy way.If you want to preserve the organisms for DNA analysis ethanol is the standard. Formalin-based and other preservatives. If you canu2019t get reagent grade ethanol, grain alcohol or even vodka can work. This would also work if you are interested in examining the frustules of diatoms, although the diatoms are often cleaned further with sulfuric acid if this is the final goal.For zooplankton with hard shell such as copepods, cladocera and fish larvae, formalin is the standard solution. Formalin is hazardous so samples are often transferred to ethanol before examination. Even when examining ethanol preserved samples, a vent hose is typically located next to the microscope to draw off the alcohol fumes. Zooplankton is typically examined in an open petri dish under a dissecting microscope. For protozoa and dinoflagellates, I used Lugolu2019s Iodine Solution. This tends not to rupture fragile cells. These organisms are typically examined in a Sedgewick-Rafter cell under a compound microscope or maybe with an inverted microscope. Since the sample is covered, there is usually not a need for extra ventilation. The safest way to preserve general plankton samples is probably with ethanol. More fragile cells will lyse and the phytoplankton will lose most of its chlorophyll, but it is the least toxic preservative available. You can filter it onto a fins mesh and rinse it with water before examining it if it is for a school project. If you only need to preserve the sample for a short period of time you can store it in an opaque jar in the refrigerator. Cold-water organisms may be able to survive for a while under these conditions, and whatever dies will decay more slowly than at room temperature. If you chose this method, only fill the sample container 1/3 of the way with water. If you fill it up all the way, the only oxygen available is what is in the water at the time. By leaving a lot of air, oxygen exchange can occur. The same goes when bagging aquarium fish. no more than 1/3 of the bag should be filled with water, the rest should be air.If you want to look up specific methods I suggest looking up the following journals online: Limnology & Oceanography, Marine Ecology Progress Series and the Journal of Plankton Research. They have made many of the older papers free for anyone to view, so if you look up plankton studies you can find out which methods people have used. The plankton are so diverse that no single method preserves everything equally well.What are some easy ways to store freshwater samples from ponds, preserving the life forms (microorganisms like plankton, algae)? Would freezing work?------3. How do we know the full 3-dimensional shape of a cell or any extremely diminute things?Cells are actually quite large when you consider that with electron microscopy, scientists can resolve structures that are less than a nanometer in one dimension. Much of what we know about intracellular structures came from early transmission electron microscopy (TEM) images of extremely thin slices of tissues. Using plastics, we can immobilize cells in a solid block and cut them into tiny slices (10s of nm thick) using extremely sharp and precise blades. We can image these thin sections of cells using heavy metals as contrasting agents. Thicker tissue sections (5-10 um) can be used in combination with non-specific tissue and nuclear stains (such as hematoxylin) and light microscopy to visualize cell shape, size, and position in a tissue. Much of what we know about the structure and shape of cells is through histological sections of tissues. Figure 1: Representative TEM image of a plant cell.Figure 2: Representative H&E stain of intestinal tissue.Phase contrast and brightfield imaging (using conventional upright or inverted microscopes equipped with medium-power objectives) enable scientists to visualize the outline and the nucleus of cells, so these techniques have long been used to look at the shape of cells in a two-dimensional context (on a tissue culture plate, for example). Microscopes with fluorescent light sources and sensitive CCD cameras enable scientists to visualize inside and outside of cells using fluorescent probes (that can bind to particular components of the cell, either on the inside of the cell or outside). Confocal laser-scanning microscopy is a very similar technique to fluorescence microscopy, but the technique uses very high power lasers to excite particular fluorophores in your cell samples through an extremely small aperture (that eliminates much of the background light characteristic of traditional fluorescence microscopy). Confocal microscopy can be used to take fluorescent 2D images of the entire thickness of a cell and reconstruct these 2D images into a 3D rendering of the cell. This technique has proved indispensible in my research!Figure 3: Representative confocal reconstruction of retina, showing retinal gangliaI hope this answers your question!References: Electron microscopy (EM) Cancer: An Introduction | We All Have Unique Brains Zeiss LSM 510 Meta Confocal Microscope How do we know the full 3-dimensional shape of a cell or any extremely diminute things?