BACKGROUND: Experimental animal models of intracerebral hemorrhag (ICH) are greatly needed, so the process of establishment should be ideal in hematoma formation and easy to operate.OBJECTIVE: To construct model of ICH in rats with double injection of autologous blood taken from the cut tail cut and double withdrawal of the needle (shortened as two-step injection model), and compare with those induced by single and double injections. DESIGN: A randomized controlled trial.SETTING: Department of Neurology, General Hospital of Military Area Command of Chinese PLA.MATERIALS: Thirty male Wistar rats of 10 to 12 months, weighing (400±25) g, provided by the Experimental Center of Medical Animals, General Hospital of Shenyang Military Area Command of Chinese PLA, were divided randomly into 3 groups with 10 rats in each group: two-step injection group, single injection group, double injection group.METHODS: The experiment was carried out in the Department of Neurology, General Hospital of Military Area Command of Chinese PLA from March to June in 2004. Autologous blood ICH model in rats were established as follows: In the two-step injection group, 50 μL unclotted autologous blood was taken from the rat tail cut, then injected with microsyringe into the caudate nucleus, 10 μL injected at first, paused for 2 minutes, and then the rest 40 μL injected slowly and continuously within 2 minutes. After the injection, the needle was kept immovable for about 4 minutes, withdrawn 2.0 mm, again kept immovable for about 4 minute, and then removed wholly at a slow speed. In the single injection group, 50 μL unclotted tail blood was injected slowly and continuously all within 2 minutes and the needle was slowly removed;④ In the double injection group, 10 mL blood was injected at first, paused for 2 minutes, the rest 40 μL injected evenly within 2 minutes, and then the needle was withdrawn slowly and uninterruptedly. Neurologic findings were scored in accordance with Longa’s five-point scale (0-4 scores, the higher the score, the severer the neurological dysfunction). The rats were killed to remove and sections were prepared, the morphological features of hematomas were grossly observed, the maximal diameter and size of hematomas in each slice were measured with the imaging analytical system, and the volume was calculated. Meanwhile, the conveniences of the techniques were compared. MAIN OUTCOME MEASURES: The morphological features and volume of hematomas, neurologic deficit score (NDS), and the convenience of the techniques were compared. RESULTS: All the 30 rats were involved in the analysis of results without deletion. ① Results of the morphological observation of volume of hematoma: In the two-step injection group, hematomas located in the right caudate nucleus area regularly in circular or analogously circular shape in each slice. The formation rate of hematomas in the single injection group and double injection group were lower than those in the two-step injection group [60% (6/10), 80% (8/10), 100% (10/10), P < 0.01, 0.05]. The volume of hematomas in the single injection group and double injection group were smaller than those in the two-step injection group [(28.5±14.8), (33.4±7.4), (41.6±3.9) mm3, P < 0.01, 0.05]. ② NDS results: The NDS scores in the single injection group and double injection group were smaller than that in the two-step injection group (0.90±0.83, 1.30±0.78, 1.90±0.57, P < 0.05). ③Comparison of the convenience of the techniques: The double injection method allowed generating reproducible hematomas in rats with shortcomings that it needed autologous arterial blood from femoral artery, and precision instruments such as microinfusion pump. The two-step injection injected fresh unclotting blood taken directly from the tail cut with microsyringe into the rat brain, and it has the advantages of easy operation, no influence on the activity of thrombase, shorter duration for model establishment, and higher rate of hematoma formation, which could generate ideal and economical models of ICH. The two-step injection induced hemotoma regularly in circular or analogously circular shape in each slice, but those induced by single and double injections were mostly in strip or fusiform shapes and extending along the needle tracks or into the ventricle or subarachnoid space. CONCLUSION: The autologous blood ICH model induced by the two-step injection method is a reproducible and reliable one in regular shape, which is better than those induced by double and single injections. BACKGROUND: Experimental animal models of intracerebral hemorrhag (ICH) are greatly needed, so the process of establishment should be ideal in hematoma formation and easy to operate. OBJECTIVE: To construct model of ICH in rats with double injection of autologous blood taken from the cut tail cut and double withdrawal of the needle (shortened as two-step injection model), and compare with those induced by single and double injections. DESIGN: A randomized controlled trial. SETTING: Department of Neurology, General Hospital of Military Area Command of Chinese PLA. MATERIALS: Thirty male Wistar rats of 10 to 12 months, weighing (400 ± 25) g, provided by the Experimental Center of Medical Animals, General Hospital of Shenyang Military Area Command of Chinese PLA, were divided into 3 groups with 10 rats in each group: two-step injection group, single injection group, double injection group. METHODS: The experiment was carried out in the Department of Neurology, General Hospital of Mili tary Area Command of Chinese PLA from March to June in 2004. Autologous blood ICH model in rats were established as follows: In the two-step injection group, 50 μL unclotted autologous blood was taken from the rat tail cut, then injected with microsyringe into the caudate nucleus, 10 μL injected at first, paused for 2 minutes, and then the rest 40 μL injected slowly and continuously within 2 minutes. After the injection, the needle was kept immovable for about 4 minutes, withdrawn 2.0 mm, again kept immovable for about 4 minute, and then removed wholly at a slow speed. In the single injection group, 50 μL unclotted tail blood was injected slowly and continuously all within 2 minutes and the needle was slowly removed; ④ In the double injection group, 10 mL blood was injected at first, paused for 2 minutes, the rest 40 μL injected balanced within 2 minutes, and then the needle was withdrawn slowly and uninterruptedly. Neurologic findings were scored in accordance with Longa’s five-po int scale (0-4 scores, the higher the score, the severer the neurological dysfunction). The rats were killed to remove and sections were prepared, the morphological features of hematomas were grossly observed, the maximal diameter and size of hematomas in each slice were measured with the imaging analytical system, and volume was calculated. MAIN OUTCOME MEASURES: The morphological features and volume of hematomas, neurologic deficit score (NDS), and the convenience of the techniques were compared Results of the morphological observation of volume of hematoma: In the two-step injection group, hematomas located in the right caudate nucleus area regularly in circular or analogously circular shape in each slice. The formation rate of hematomas in the single injection group and double injection group were lower than those in the two-step injectio The volume of hematomas in the single injection group and double injection group were smaller than that of the control group (n group [60% (6/10), 80% (8/10), 100% those in the two-step injection group [(28.5 ± 14.8), (33.4 ± 7.4), (41.6 ± 3.9) mm3, P <0.01, group were smaller than that in the two-step injection group (0.90 ± 0.83, 1.30 ± 0.78, 1.90 ± 0.57, P <0.05). ③Comparison of the convenience of the techniques: The double injection method allowed generating reproducible hematomas in rats with shortcomings that it needed autologous arterial blood from femoral artery, and precision instruments such as microinfusion pump. The two-step injected injected fresh unclotting blood taken directly from the tail cut with microsyringe into the rat brain, and it has has advantages of easy operation, no influence on the activity of thrombase, shorter duration for model establishment, and higher rate of hemato ma formation, which could generate ideal and economical models of ICH. The two-step injection induced hemotoma regularly in circular or analogously circular shape in each slice, but those induced by single and double injections were mostly in strip or fusiform shapes and extending along the needle tracks or into the ventricle or subarachnoid space. CONCLUSION: The autologous blood ICH model induced by the two-step injection method is reproducible and reliable one in regular shape, which is better than those induced by double and single injections.