吉姆萨色素 (又称姬姆萨色素) 是由天青II与伊红混合而成，Giemsa染色原理和结果与瑞氏染色基本相同，吉姆萨染色对胞浆着色力较强，能较好的显示胞浆的嗜碱性程度，特别是对血液和骨髓细胞中的嗜天青、嗜酸性、嗜碱性颗粒，着色清晰，但是对胞核着色偏深,核结构显色不佳，故吉姆萨染液常与瑞氏染液联合使用。许多染料对DNA都有不同程度的亲和性，故都可用作染色体的染色，在染色体的常规染色中一般用吉姆萨、地衣红、福尔根、石碳酸复红等都可获得良好的染色效果，G显带技术是指将染色体玻片标本经过胰蛋白酶处理后，再用吉姆萨进行染色，使每条染色体沿其长轴显示出一定数量的、宽窄和深浅不同的横纹，即带型，由于人类22种常染色体和X、Y染色体的带型各具特征，根据带型可清楚地分辨出每条染色体。
阿拉丁吉姆萨染色液 (染色体专用) 由10×储存液和磷酸盐缓冲液组成，1:9混合后使用，主要用于染色体G带染色，以进口的姬姆萨色素、甲醇为主要原料，含特有衬染剂，经研磨配制而成，能呈现出清晰的细胞染色效果，经常用于组织切片、血液和细胞涂片、细菌、染色体显带、原生动物寄生虫等染色。该试剂仅用于科研领域，不适用于临床诊断或其他用途。
产品组分及储存条件：
J1510432
| Component
| 100mL
| 500mL
| Storage
|
J1510432A
| Giemsa Stain 储存液(10x)
| 10mL
| 50mL
| RT
|
J1510432B
| 磷酸盐缓冲液
| 100mL
| 500mL
| RT |
自备材料：
1、PHA、秋水仙素、0.075M氯化钾溶液、甲醇乙酸固定液(甲醇：冰乙酸=3：1)
2、恒温培养箱、载玻片、显微镜
操作步骤(仅供参考)：
1、配制 Giemsa Stain 工作液：按试剂(A)：试剂(B)=1：9混合，即取1份 Giemsa Stain 储存液 (10×) 加入到9份的磷酸盐缓冲液中充分混匀，即为 Giemsa Stain 工作液，该工作液为即用型试剂，不易保存，即用即配；如效果不佳，可改变配制比例。
2、培养：取人类外周血3~5ml在PHA (一般工作浓度约60μg/ml)、血清等条件下体外培养3天，加入BrdU和(或)秋水仙素 (一般工作浓度约0.5μg/ml)，培养30~60min并收集细胞即2000g离心5min，留取沉淀。
3、低渗：加入5~8ml提前37°C预热的0.075M氯化钾溶液37°C低渗处理20~30min。
4、固定：加入1ml新配制的甲醇乙酸固定液，轻轻混匀，2000g离心5min，留取沉淀；再次加入2~3ml甲醇乙酸固定液，轻轻混匀，室温固定15~20min，2000g离心5min，留取沉淀；再次加入2~3ml甲醇乙酸固定液，重复该步骤，留取沉淀。
5、滴片：加入少许甲醇乙酸固定液至沉淀中，轻轻混匀制成细胞悬液，滴加2~3滴细胞悬液于提前遇冷的载玻片上，吹散水蒸气熏蒸促使细胞破裂 (亦可高空垂直滴下细胞悬液促使细胞破裂)，70°C烤片2h。
6、消化：用0.025~0.05%胰蛋白酶溶液处理载玻片30~90min，依据胰蛋白酶的浓度、批次等摸索处理时间。
7、染色：滴加 Giemsa Stain 工作液覆盖玻片，室温染色15～30min。
8、用自来水或蒸馏水缓慢从载玻片一端冲洗，将多余染色液冲掉，空气干燥。
9、先用低倍镜浏览整张玻片标本，找到分散良好且染色体长短适中的分裂相，再用油镜观察并识别染色体G显带核型。
注意事项：
1、涂片染色中Giemsa染色后请勿先去除染液或直接对涂片用力冲洗。
2、如果染色过深或过浅，应调整染色时间或工作液浓度。
3、pH值对染色有一定影响，载玻片应清洁、无酸碱污染，以免影响染色效果。
4、染色液经稀释后液面有金属光泽则表示染液有染色作用，否则染色液可能失效。
5、染色液可重复使用，但不能多次重复，若有沉淀物应过滤后使用。
Giemsa stain (also known as Gimsa stain) is a mixture of Azure II and Eosin. The staining principle and results of Giemsa staining are basically the same as those of Wright staining. Giemsa stain has a strong staining affinity for cytoplasm, and can well demonstrate the degree of cytoplasmic basophilia. In particular, it can clearly stain the azurophilic, eosinophilic and basophilic granules in blood and bone marrow cells. However, it stains cell nuclei too deeply, resulting in poor visualization of nuclear structures. Therefore, Giemsa stain solution is often used in combination with Wright stain solution. Many dyes have varying degrees of affinity for DNA and thus can be used for chromosome staining. For routine chromosome staining, satisfactory results can generally be obtained with Giemsa stain, Orcein stain, Feulgen stain, Carbol Fuchsin stain and other dyes. G-banding technology refers to treating chromosome slide specimens with trypsin, followed by staining with Giemsa stain. This process makes each chromosome display a certain number of horizontal bands with different widths and shades along its long axis, which are called banding patterns. Since the 22 types of human autosomes and the X and Y sex chromosomes each have distinct banding patterns, each chromosome can be clearly identified based on its banding pattern.
Aladdin Giemsa Stain Solution (for Chromosomes) consists of a 10× stock solution and phosphate buffer solution, which should be mixed at a ratio of 1:9 before use. It is mainly used for chromosome G-banding staining. The solution is prepared by grinding and mixing, with imported Giemsa stain and methanol as the main raw materials, plus a proprietary counterstain. It can produce clear cell staining results, and is commonly used for staining tissue sections, blood and cell smears, bacteria, chromosome banding, protozoan parasites, etc. This reagent is for research use only and is not suitable for clinical diagnosis or other purposes.
Product Components and Storage Conditions:
J1510432
| Component
| 100mL
| 500mL
| Storage
|
J1510432A
| Giemsa Stain Stock Solution (10×)
| 10mL
| 50mL
| RT
|
J1510432B
| Phosphate Buffer Solution
| 100mL
| 500mL
| RT |
Self-prepared Materials:
1. PHA, colchicine, 0.075M potassium chloride solution, methanol-glacial acetic acid fixative (methanol:glacial acetic acid = 3:1)
2. Constant temperature incubator, glass slides, microscopes
Operating Procedures (For Reference Only):
1. Prepare Giemsa Stain Working Solution: Mix Reagent (A) and Reagent (B) at a ratio of 1:9. Specifically, add 1 part of 10× Giemsa Stain Stock Solution to 9 parts of phosphate buffer solution and mix thoroughly to obtain the Giemsa Stain Working Solution. This working solution is ready-to-use, not suitable for long-term storage, and should be freshly prepared before use. If the staining effect is unsatisfactory, the mixing ratio can be adjusted accordingly.
2. Cell Culture: Collect 3–5 ml of human peripheral blood and culture it in vitro for 3 days under conditions supplemented with PHA (general working concentration: approx. 60 μg/ml) and serum. Add BrdU and/or colchicine (general working concentration: approx. 0.5 μg/ml), continue culturing for 30–60 min, then harvest the cells by centrifugation at 2000 g for 5 min and retain the cell pellet.
3. Hypotonic Treatment: Add 5–8 ml of pre-warmed 0.075 M potassium chloride solution (pre-heated to 37°C) to the cell pellet, incubate at 37°C for 20–30 min for hypotonic treatment.
4. Cell Fixation: Add 1 ml of freshly prepared methanol-glacial acetic acid fixative to the pellet, mix gently, centrifuge at 2000 g for 5 min and retain the pellet. Add another 2–3 ml of methanol-glacial acetic acid fixative, mix gently, fix at room temperature for 15–20 min, centrifuge at 2000 g for 5 min and retain the pellet. Repeat this fixation step with another 2–3 ml of methanol-glacial acetic acid fixative and retain the final pellet.
5. Slide Dropping: Add a small amount of methanol-glacial acetic acid fixative to the cell pellet and mix gently to prepare a cell suspension. Drop 2–3 drops of the suspension onto pre-chilled glass slides. Blow away the water vapor fumes to promote cell rupture (alternatively, drop the cell suspension vertically from a height to induce cell rupture). Bake the slides at 70°C for 2 hours.
6. Trypsin Digestion: Treat the baked slides with 0.025–0.05% trypsin solution for 30–90 min. The optimal digestion time should be determined empirically according to the concentration and batch of trypsin used.
7. Staining: Add sufficient Giemsa Stain Working Solution to cover the slide surface and stain at room temperature for 15–30 min.
8. Rinsing and Drying: Rinse the slide slowly from one end with tap water or distilled water to wash off excess staining solution, then air-dry the slide naturally.
9. Microscopic Examination: First, scan the entire slide under a low-power microscope to locate well-spread metaphase cells with chromosomes of moderate length. Then switch to an oil immersion lens to observe and identify the G-banding karyotype of the chromosomes.
Precautions:
1. In the smear staining process, do not remove the stain first or rinse the smear vigorously directly after Giemsa staining.
2. If the staining is too dark or too light, adjust the staining time or the concentration of the working solution accordingly.
3. pH value has a certain impact on staining. Glass slides should be clean and free of acid-alkali contamination to avoid affecting the staining effect.
4. If the diluted stain shows a metallic luster on the liquid surface, it indicates that the stain is effective; otherwise, the stain may have expired.
5. The stain can be reused, but not for multiple times. If there is any sediment, filter the stain before use.
ZJ26F0433808
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