This experiment is mainly used for routine blood tests.
Operation method
microscopic examination of thin section of specimen as part of biopsy
Principle
There are two components in the Riesling dye, merocyanine and eosin, the former being basic and the latter acidic, which have different affinities for various substances in the cells, causing them to show different shades of color. The nucleus of the blood cell consists of deoxyribonucleic acid and the strongly basic histones, arginins, etc., which form nucleoproteins. This strong alkaline material and Ruishi dye in the acidic dye red have affinity, so stained red; nucleoprotein and a small amount of weak acidic protein, they and the basic dye in the dye solution of the role of blue and stained blue, but the content is too small, the blue reaction is very weak, so the nuclear staining showed purplish-red. The cytoplasm of naive cells and the nucleus of the nucleolus contain acidic substances, which have an affinity with the basic dye in the dye, so it stains blue.
Materials and Instruments
Blood Move 1. Preparation of blood smear (1) Place a small drop of blood (about 5 ul) on a clean slide and push the slide at an angle of about 30°, with even and rapid pressure and without repetition. (2) If the drop of blood is small, push the slide more slowly and at an angle of less than 30° to produce a thinner smear. (3) If the droplet is large, the push is fast, and the angle is greater than 30°, the smear is thicker. 2. Place the specimen on a flat surface (preferably on a fixed frame) and apply the dye to the smear with a dropper, which can be used to disperse the dye to cover the whole smear. 3. After the staining solution covers the whole smear, wait for a moment or add buffer solution immediately and make the buffer solution mix well with the staining solution. 4. Ratio of dye solution to buffer solution (1) The amount of dye solution should be sufficient, otherwise the dye solution will evaporate quickly and precipitate the dye on the cells. (2) The ratio of dye solution to buffer is about 1:2~4 which is more appropriate. (3) The greater the dilution, the longer the dyeing time, the better the cell coloring, and vice versa. (5) Staining time: depends on the specific situation, generally takes about 10~30 minutes. 6. Rinsing: Rinse the dye on the smear with tap water. 7. 7. Microscopic examination: After natural drying, use a light microscope to examine. Caveat 1. Blood smears should be made to avoid air bubbles and be of moderate thickness. 2. The ratio of dye to buffer should be moderate. Common Problems I. Normal blood cell morphology 1. Mature erythrocytes: Normal erythrocytes are slightly concave on both sides and appear as discs; after staining, they appear as orange-red cells with light staining in the center, with an average diameter of 7.6 um. For more product details, please visit Aladdin Scientific website.
Riesling's dye Pure methanol Potassium dihydrogen phosphate Disodium hydrogen phosphate
Microscope Slide Coverslip Dropper
2. Mature neutrophils: there are two types of neutrophils in PB, rod-shaped nuclei and lobulated nuclei. Rod-shaped nucleus granulocyte morphology: round or oval, diameter 10~13 um. nucleus such as rods, ribbons, the nucleus of the concave more than assumed that the nucleus of the round diameter of 3/4, and the two ends of the inside and outside of a considerable period of parallel. Chromatin is clumped and unevenly colored, and there may be blank areas in between. The cytoplasm no longer contains basophilic material and is filled with neutral granules. Neutral lobulated nucleated granulocytes: round, 10-13 um in diameter, the nucleus is lobulated, with two lobes in a few cases, and six or seven lobes in many cases, with thin filaments connecting or completely disconnecting the lobes, or overlapping each other. Chromatin was finely clumped. The cytoplasm is covered with neutral granules.
3. Mature lymphocytes: round, 5-18 um in diameter, with rounded or fava bean-like nuclei and dense, clumped chromatin. The cytoplasm is small, blue, and may have a few round, peripheral, neatly organized azurophilic granules. Large lymphocytes have a large amount of cytoplasm and are light blue in color.
4. Monocytes: round or irregular, 12-20 um in diameter, with centered or lateralized nuclei and irregular shapes. Chromatin was agglutinated like a sieve bottom or a coarse line of reticulation, but there were no agglutinated masses of chromatin. The cytoplasm was abundant, gray-blue in color, scattered with many tiny pink granules, and the granules were so numerous that the cytoplasm became pink, and vacuoles and exoplasma could be present.
5. Platelets: Normal platelets are star-shaped, comma-shaped or irregular, with a diameter of 2~5 um, light blue in color, with a number of small light purplish-red granules in the center.