This suggests that inbred mice are more sensitive
There have been clear international Laboratory Consumables regulations for experimental animals. Otherwise, mice are prone to death.) cannot be transported under normal circumstances. Adding litter and diet to the mouse box 4. Cooperation between various scientific research units Increasing exchanges have increased the demand for mouse transportation, including model mice customized by third-party institutions, animals donated by scientific research partners, and even experimental animals from abroad.
Cardboard box, if you need to transport it for a long time, you need to choose a wire box or plastic box lined with wall paper or calcium plastic. To check the time it takes for animal weight to recover from transport stress, the researchers measured the weight of mice and rats at the same age and described the changes in body weight.
This suggests that inbred mice are more sensitive to stress than closed-group mice. References:  A report from the American Veterinary Medical Association Animal Air Transportation Study Group. People have not paid enough attention to the phenomenon that the stress response of animals during transportation affects the experimental results. While providing sufficient airflow, it can also prevent the entry of airborne particles and affect the health of mice. In such cases, mice need to be transported to other cities nearby by air, and the cost will be higher. As one of the most important model organisms, mice have been in increasing demand in recent years. In general, short-distance transportation in the city can choose to line the fine wire mesh.), the transportation time is about 24 hours (including self-packed small Rat-time for unpacking)
, the average weight of the mice will be reduced by about 0. 2012 29 (3): 15-19. If the truck is arranged to be transported by land, it is best to keep the temperature inside the car at about 25 ° C and pay attention to ventilation and ventilation. If the transportation time is long, the distance is long, the packing density and weight consumption may reach 10-15%, requires attention. Listen to the lab's brothers and sisters saying that it is now over 30 degrees high, and you can sweat for a few minutes outside.
Centrifuge again and try to aspirate all the liquid
Carefully aspirate the formalin. Prior to tubes factory the sections were placed in xylene. If blocky matrigel is observed, incubate for 30 minutes on ice using BD cell recovery medium. Mix gently and incubate on ice for at least one hour. Block with a suitable blocking buffer at room temperature for 1 hour (or block according to the usual blocking method). Centrifuge again and try to aspirate all the liquid.05% tween-20, pH 6. hPSC-Derived Intestinal Organoid, EPCAM (Green), KRT20 (Red), Desmin (White), DAPI (Blue) 8. Rinse with distilled water. 5.
Staining (Alsin Blue) a) 3% acetic acid solution: 3 mL of glacial acetic acid, dH2O 97 mLb) Alcian Blue solution (pH 2. Gently aspirate the formalin without touching the precipitated organoids.C. Antigen retrieval 7. Dissolve the dye in distilled water and stir with acid. If using a 200 micro; L or smaller tip, cut off the tip and tail to prevent damage to the organoid. Prepare the mold: prepare an 8-strip PCR tube, divide it into individual tubes, and cut off the lower half of the tube To get a ring as a mold.T. 10. Note: In terms of staining, organoid sections are treated in the same way as other tissue sections, using the usual steps of general dewaxing, fluid replacement, and antigen retrieval. 12. Adjust the pH with acetic acid solution and filter *.
Use a 200 micro; L pipette tip to gently squeeze the coverslip to squeeze out air bubbles. Prepare 1N hydrochloric acid: 915 mL of distilled water, 85 mL of concentrated hydrochloric acid, Alcian Blue, pH 0. Fix the sections with mounting medium and seal with nail polish. Dry the section and mark the section of the organoid section with an immunohistochemical pen. Steps 11 and 12 must be completed as quickly as possible (within 15 seconds), otherwise the agarose will solidify in the pipette tip and cause sample loss. Wash twice with PBS and once with dH2O. (To help remove Matrigel, you can also add Cell recovery medium and incubate on ice for 15-30 minutes. Place sections in 3% acetic acid solution for 3 minutes (used as mordant). 6.
This step needs to be done in a fume hood. Note:-Immunofluorescent staining, please proceed to steps 7-18-H amp; E staining, please proceed to steps 19-24-Alcian Blue staining,
This method is also the mainstay in the field of tumor
In the first few issues, we have learned the basic characteristics and culture methods of tumor cells. Next, we will introduce various commonly used cell experimental techniques. For any research, the logical thinking of a system seems to be more important than the specific experimental method. So before the introduction, we will talk about the research ideas of previous people and some common methods and techniques in oncology research from the perspective of drug development. Welcome to discuss each other. Part 1: Drug screening mode represented by NCI This screening mode is the NCI-60-based anti-tumor drug screening mode. Since 1990, the industry and academia have screened more than 100,000 compounds using NCI-60 cells.
This method is also the mainstay in the field of tumor drug discovery. First look at the four stages of its development: 1955-1985: in vivo screening model. Based on the transplanted tumor animal model, based on the animal's life extension rate and tumor growth inhibition rate as the basic evaluation indicators of the drug; 1985-1990: in vitro screening model (in vitro) gradually replaced the in vivo model. That is to say, abandon in vivo mouse screening and an in vitro antitumor drug screening model based on human tumor cell lines. 1990-2016: Based on an in vitro screening model.
Mainly in vitro screening of nine major categories of 60 human tumor cell lines NCI-60; subsequent verification in vivo ; 2016-present: the use of patient-derived xenograft tumors (PDX) as a model. It is mainly considered that NCI-60 has been cultured for thousands of generations in vitro and may have mutated. However, the establishment of the PDX model is difficult and the database is not complete. At present, we still adopt a strategy based on in vitro cell line screening and subsequent verification of the in vivo model. The basic screening model is shown in the following figure: NCI-60-based drug screening process [1
] Drug screening process (screening based on cell phenotype): First screening of compounds to be screened (3 sensitive tumor cell lines MCF-7; NCI -H460; SF-268), with inhibition of cell proliferation as an indicator (3 parameters: semi-growth inhibiting GI50, overall growth-inhibiting TGI and LC50). Further screening on 15-20 tumor cell lines (including drug-resistant cell lines such as MCF-7 / ADR), normal cell lines, etc .; comprehensive in vitro results, validation in vivo models; drug safety, pharmacokinetics, etc. (in vivo Animal level);