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致畸试验骨组织染色液 (茜素红应用液Ⅰ)
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概述
致畸试验骨组织染色液 (茜素红应用液Ⅰ) 为预配完成的即用型骨骼染色工作液，由茜素红、冰乙酸、甘油、水合氯醛、氢氧化钾复配制成，液体呈均匀红棕色，具备轻微刺激性与弱腐蚀性。适用于食品、化妆品、保健用品、工业化学品毒理学安全性评价，专供大鼠、小鼠、家兔致畸试验中经氢氧化钾透明后的胎仔整体骨骼染色检测。
染色原理：试剂中茜素红可特异性与胎仔骨骼钙盐发生螯合反应，生成稳定紫红色复合物，完整标记头骨、肋骨、胸骨、脊柱、四肢骨的骨化中心、骨缝结构，清晰识别骨缺损、骨骼融合、肋骨分叉、骨化不全等各类致畸相关骨骼发育畸形。
该试剂仅用于科研领域，不适用于临床诊断或其他用途。
操作步骤(仅供参考)：
1、样本预处理(大鼠、小鼠适用)：
1)取每窝 1/2 剖宫获取的活胎仔，95%乙醇密封固定 2-3 周；
2)流水冲净固定残留，放入致畸试验骨组织透明液(KOH 溶液)(B1529929)浸泡 8-72h，直至软组织半透明、完整骨骼轮廓清晰，操作轻柔防止肋骨、脊柱断裂。
注：家兔致畸试验不可分半取样，全部胎仔均需完整染色检查。
2、骨骼染色(本品直接使用，无需稀释)：
1)将完成透明处理的胎仔完全浸没于致畸试验骨组织染色液 (茜素红应用液Ⅰ)(B1529924)中，容器密封避光、室温静置染色 6-48h；
2)每日轻摇容器 1-2 次，直至头骨、四肢骨均匀呈紫红色即可，无需过度延长染色时间。
3、分级透明脱色(配套试剂需自备)：
1)染色结束后取出胎仔，浸入致畸试验骨组织透明液 A(KOH-甘油溶液)(B1529923)避光浸泡 1-2d；
2)转入致畸试验骨组织透明液 B(甘油溶液)(B1529922)避光浸泡 2-3d；
3)若软组织紫红色残留较重，可适度延长透明处理时长。
4、镜检观察与统计规范
1)标本脱色完成后转移至纯甘油中长期封存，方便镜检与标本留存；
2)采用透射光源，体视显微镜下依次检查颅骨、脊柱、肋骨、胸骨、四肢骨，记录骨化缺失、骨骼融合、肋骨分叉、骨缺损、骨化发育不全等各类骨骼畸形；
3)毒理试验统计要求：所有骨骼畸形、发育异常指标均以窝为单位进行统计，计算总畸胎率、单项骨骼畸形率。
注意事项：
1、本品为预配应用液，已添加氢氧化钾，禁止额外加水、酸碱、强碱原液稀释，否则染料析出、染色失效。
2、染色时长可根据胎鼠日龄、标本大小灵活微调，避免染色过深导致软组织背景发红，干扰畸形判断。
3、染色时间需根据标本大小和发育阶段灵活调整，避免染色过深或过浅影响观察。
4、仅适用于经 KOH 透明后的完整胎仔整体骨骼染色，不适用于石蜡切片、细胞钙化染色、脱钙组织染色。
5、为了您的安全和健康，请穿实验服并戴一次性手套操作。
Bone Tissue Staining Solution (Alizarin Red Working Solution I) for Teratogenicity Testing is a ready-to-use skeletal staining working solution. It is formulated with Alizarin Red, glacial acetic acid, glycerol, chloral hydrate, and potassium hydroxide. The solution is uniformly reddish-brown and exhibits slight irritancy and weak corrosiveness. It is suitable for toxicological safety evaluation of food, cosmetics, health products, and industrial chemicals, and is specifically intended for whole-mount skeletal staining of fetuses from rats, mice, and rabbits after potassium hydroxide clearing in teratogenicity studies.
Staining Principle: Alizarin Red in the reagent specifically chelates with calcium salts in fetal skeletal tissue to form a stable purplish-red complex. This allows clear visualization of ossification centers and suture structures in the skull, ribs, sternum, vertebral column, and limb bones, facilitating identification of teratogenicity-related skeletal developmental abnormalities such as bone defects, skeletal fusion, bifurcated ribs, and incomplete ossification.
This reagent is intended for research use only and is not suitable for clinical diagnosis or other purposes.
Operating Procedure (For Reference Only):
1. Sample Pretreatment (Applicable to Rats and Mice):
1) Select 1/2 of the live fetuses obtained by cesarean section from each litter and fix them in 95% ethanol in a sealed container for 2-3 weeks;
2) Rinse thoroughly under running water to remove residual fixative, then immerse the fetuses in Bone Tissue Clearing Solution (KOH Solution) for Teratogenicity Testing (B1529929) for 8-72 h until the soft tissues become translucent and the complete skeletal contours are clearly visible. Handle gently to avoid fractures of the ribs and vertebral column.
Note: For rabbit teratogenicity tests, half-litter sampling is not applicable. All fetuses must undergo complete skeletal staining and examination.
2. Skeletal Staining (Use Directly Without Dilution):
1) Completely immerse the cleared fetuses in Bone Tissue Staining Solution (Alizarin Red Working Solution I) for Teratogenicity Testing (B1529924), seal the container, protect from light, and allow to stain at room temperature for 6-48 h;
2) Gently shake the container 1-2 times daily until the skull and limb bones are uniformly stained purplish-red. Excessive extension of the staining time is unnecessary.
3. Graded Clearing and Decolorization (Supporting Reagents Required):
1) After staining, remove the fetuses and immerse them in Bone Tissue Clearing Solution A (KOH-Glycerol Solution) for Teratogenicity Testing (B1529923), protected from light, for 1-2 d;
2) Transfer the fetuses to Bone Tissue Clearing Solution B (Glycerol Solution) for Teratogenicity Testing (B1529922), protected from light, for 2-3 d;
3) If substantial purplish-red coloration remains in the soft tissues, appropriately extend the clearing time.
4. Microscopic Examination and Statistical Analysis
1) After decolorization is complete, transfer the specimens to pure glycerol for long-term preservation to facilitate microscopic examination and specimen storage;
2) Using transmitted light, examine the skull, vertebral column, ribs, sternum, and limb bones sequentially under a stereomicroscope, and record skeletal abnormalities including absent ossification, skeletal fusion, bifurcated ribs, bone defects, and incomplete ossification;
3) Toxicological test statistics: All skeletal malformations and developmental abnormalities should be analyzed on a litter basis, and the total fetal malformation rate and individual skeletal malformation rates should be calculated.
Precautions:
1. This product is a ready-to-use working solution containing potassium hydroxide. Do not further dilute it with water, acids, alkalis, or concentrated alkaline solutions, as this may cause dye precipitation and loss of staining performance.
2. The staining time may be adjusted according to the age of the fetus and specimen size. Avoid overstaining, which may cause excessive red background staining of soft tissues and interfere with the evaluation of skeletal abnormalities.
3. The staining duration should be adjusted according to specimen size and developmental stage to avoid excessively strong or weak staining that may affect observation.
4. This product is intended only for whole-mount skeletal staining of intact fetuses after KOH clearing. It is not suitable for paraffin sections, cellular calcification staining, or staining of decalcified tissues.
5. For your safety and health, wear a laboratory coat and disposable gloves during operation.
规格
规格或纯度
组织学级, BioReagent, 生物染色剂
稳定性与储存
Store at room temperature long term (12 months).
英文名称
Bone Tissue Staining Solution (Alizarin Red Working Solution I) for Teratogenicity Testing
储存条件
室温
运输条件
常规运输
产品线
生物产品
名称和识别符
分子类型
生物试剂/缓冲液
高级数据
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