top student at medical school

Chapter 68 Huge Profits (Please add to your favorites and read on!)

The National High Technology Research and Development Program (863 Program) and the National Key Basic Research Program (973 Program, including the Major Scientific Research Program) are both relatively high-end national research projects.

To obtain any one of them, sufficient accumulation is required.

Of course, securing even one can bring great success and fame. A few million in research funding is a piece of cake, and tens of millions in research grants are quite common.

The initial funding for such a project is more than a hundred times that of the youth grant that Yuan Weihong has secured so far.

Just as I was about to say something, the patient who had finished his check-up and follow-up appointment outside opened the door.

Yuan Weihong stopped talking, and Fang Ziye quickly began to gather his knowledge points, examining the images step by step, making accurate diagnoses and prescribing medications, and then obtained 0.9 knowledge points.

They felt a surge of joy and happiness.

Even Zhongnan Hospital isn't the kind of hospital that deals with all sorts of difficult and complicated cases. While it certainly has its share of such cases, it also has some simpler ones. When patients register, they don't consider the idea of ​​wasting resources.

After finishing his outpatient visit, Fang Ziye gained about 15 more knowledge points. This was enough to upgrade his knot-tying skills to level 3 and give gifts to his teacher.

Fang Ziye's knowledge points balance has suddenly accumulated to over 60. He can add another level 3 skill, and tonight he also has a cell experiment that will soon yield results. When he observes the results, he will definitely gain another considerable amount of knowledge points.

"Master, I'll put my lab coat down first, then I'll go to the lab!"

“Okay, WB, run well.” Yuan Weihong instructed, as if giving Fang Ziye a hint, indicating that he knew exactly what Fang Ziye was doing.

Fang Ziye nodded and quickened his pace.

Conducting experiments, raising funds, publishing articles, and acquiring knowledge—it's fulfilling and enjoyable.

WB is an abbreviation for Western blotting, which is a method of transferring proteins onto a membrane and then detecting them using antibodies.

The ultimate result of gene expression is the production of the corresponding protein (or enzyme). The HK2 studied by Fang Ziye is actually the first rate-limiting enzyme in the biochemical glycolysis process, hexokinase. HK2 is an abbreviation for hexokinase. Hexokinase is an enzyme that catalyzes the phosphorylation of hexoses. It is the first enzyme in the glycolysis pathway and also the absolute rate-limiting enzyme in the glycolysis pathway.

As we all know, glycolysis is the first step in converting sugar into energy. Without this first step of glycolysis, it would be almost impossible for glucose and fructose to be used normally. And relying on ketone bodies or other alternative pathways as substrates for aerobic oxidation would certainly be of no use.

Therefore, quantitative detection of HK2 levels, specifically the absolute amount after overexpression and silencing in giant cell tumors of bone, provides quantitative evidence for the role of HK2 in the development of giant cell tumors of bone.

Western blotting uses polyacrylamide gel electrophoresis. The analyte is protein, the probe is antibody, and the colorimetric assay is performed using labeled secondary antibody.

Protein samples separated by PAGE are transferred to a solid support (e.g., a nitrocellulose membrane). The solid support adsorbs proteins non-covalently, preserving the types and biological activities of the peptides separated by electrophoresis. The proteins or peptides on the solid support serve as antigens, triggering an immunoreaction with corresponding antibodies, followed by a reaction with enzyme- or isotope-labeled secondary antibodies. Substrate staining or autoradiography is then used to detect the protein components expressing the specific target gene separated by electrophoresis…

The principle sounds quite complicated, but in practice, the main steps are as follows.

The reagent preparation, which is essentially the track for proteins to run, requires polyacrylamide gel electrophoresis.

Then there are the athletes participating in the competition. After lysing the tumor cells, the internal proteins were fully released, and then a standard curve was created.

After creating the standard curve, you can start training the 'athletes' produced by the cells you've processed.

Following a series of processes including electrophoresis, transfer, immunoassay, and development, the target protein is visualized on a gel film using specialized antibodies and reagents. The film is then scanned or photographed, and a gel imaging system is used to analyze the molecular weight and net optical density of the target band…

This process is complex and relatively long, requiring a lot of patience.

However, since Fang Ziye had already finished his outpatient clinic and was in the lab, he proceeded step by step without any problems. Luo Tingzhu was also there to help him out.

After the final result was presented, Luo Tingzhu was overjoyed: "Senior Brother Fang, your band is very clear today. Last time, Senior Brother Fan's band was dumbbell-shaped."

After she finished speaking, she turned her head to look at Fang Ziye's finished product. The stripes were clear, there were no stray stripes, and the background was clean!

"A dumbbell shape usually indicates a problem with the gel preparation. Another possibility is that the sample contains too many impurities that were not centrifuged, causing the impurities to deposit in the middle of the wells, while the protein is naturally pushed to the sides."

"Prepare the glue properly, and never use any that is substandard. Don't take any risks." Fang Ziye carefully considered the principle and explained it to Luo Tingzhu in this way.

"What if there's a trailing stripe?" Luo Tingzhu thought for a moment and then asked another question, probably because she had seen this situation when she was doing it with others.

"If the bands are trailing, it usually means that the protein concentration is high or the incubation time and concentration of the primary antibody are too long."

“If the protein content is high, it may be because the literature preparation in the early stage was not well determined. I think the most likely reason is that the primary antibody concentration is too high and the reaction time is too long.”

"Of course, there are other reasons as well." Fang Ziye looked at his finished film again.

The two bands for HK2 and β-actin (internal control) were divided into three groups: overexpression, normal expression, and silence.

For internal controls in tumor cell experiments, especially those involving the glycolysis pathway, the commonly used GAPDH (glyceraldehyde-3-phosphate dehydrogenase) cannot be used.

"You can use Prism to create this image directly, right?" Luo Tingzhu continued to ask.

"It should be about right. The difference in thickness and grayscale of the strips is quite obvious. Let's take a picture, let's take a picture..." Fang Ziye quickly asked Luo Tingzhu to help take a picture as well.

Luo Tingzhu readily agreed, as lively as a diligent little squirrel.

At this moment, Fang Ziye watched as his knowledge points rapidly jumped to 134, an increase of a full seventy points. He knew that this WB experiment played a significant role in this project, possibly serving as the small framework supporting the entire project.

"How was the photo?" Luo Tingzhu asked Fang Ziye after taking the picture.

Fang Ziye smiled like an aunt, while Luo Tingzhu thought her photography skills were pretty good, so she snapped a few more pictures.

Then comes cleaning the lab bench and washing the corresponding experimental equipment.

After everything was over, Fang Ziye suggested, "Junior Sister Luo, you've worked hard. Let's go get some late-night snacks together?"

Luo Tingzhu was flipping through the photos on Weibo, tilting her head and clicking: "Okay."

……

(End of this chapter)