新型载水飞蓟宾热敏脂质体-微泡Sono Vue复合体制备及质量分析

目的制备载水飞蓟宾热敏脂质体-微泡SonoVue复合体并进行质量分析。方法以薄膜旋蒸法制备载水飞蓟宾热敏脂质体,然后利用戊二醛偶联法将超声微泡SonoVue与载药脂质体相连接,通过紫外分光光度法检测热敏脂质体药物包封率及不同温度下释药特性。通过激光共聚焦显微镜检测载药脂质体与微泡的偶联率。结果载水飞蓟宾热敏脂质体-微泡SonoVue复合体粒径(2 724±559.9)nm,药物包封率(71.0±8.3)%,41℃时药物平均释放率明显升高,达到(55.6±7.6)%。偶联后,载药脂质体在微泡周围聚集,热敏脂质体与微泡偶联率为(90.0±4.9)%。结论载水飞蓟素热敏脂质体-微泡SonoVue复合体具有良好的偶联率、较高的药物包封率及控温释药特性,为药物递送提供一种新的载体。     

不同转染方法对荧光蛋白质粒转染人胚肾293T细胞影响的实验研究

目的:探讨荧光蛋白质粒转染人胚肾293T细胞的最佳转染方法.方法:以脂质体转染法、超声辐照法、微泡造影剂SonoVue联合超声辐照及脂质体中加入微泡造影剂SonoVue并予以超声辐照4种转染方法,将含有绿色荧光蛋白报告基因的真核表达质粒pEGFP-N1转染人胚肾293T细胞,48 h后以荧光显微镜观察293T细胞中的绿色荧光蛋白表达情况并用流式细胞仪测算转染率.结果:脂质体 SonoVue 超声辐照法的转染效果最好(23.10±2.11%).脂质体 SonoVue 超声辐照法的转染率分别与脂质体转染法、超声辐照法、微泡造影剂SonoVue联合超声辐照法的转染率比较均有显著差异(P<0.01).结论:脂质体中加入微泡造影剂SonoVue并予以超声辐照能显著提高荧光蛋白质粒在人胚肾293T细胞中的转染效果.     

超声辐照联合微泡造影剂介导人血管生成素-1基因体外转染293T细胞的研究

目的 探讨超声辐照联合微泡造影剂的基因转染系统介导人血管生成素-1(hAng-1)基因转染人胚肾293T细胞的转染效率.方法 将六孔板中的293T细胞悬液分为4组:A组,超声+质粒组,每孔加入10 μg目的 基因质粒;B组,超声+微泡+质粒组,每孔加入微泡和质粒混合液,终浓度分别为质粒10 μg/孔,微泡200 μl/孔;C组,脂质体+质粒组,每孔加入4 μl脂质体和5 μg质粒;D组,对照组,每孔仅加入10 μg目的 基因质粒.A组和B组均接受超声辐照,照射条件为连续波,声强1.5 W/cm2,作用时间为30 s.转染后48 h用荧光显微镜和流式细胞仪分别定性、定量观察基因的转染效率.结果 48 h后,荧光显微镜下观察到转染成功的细胞胞浆内发出绿色荧光,流式细胞仪检测A组、B组、C组、D组的基因转染效率分别为(3.5±0.4)%、(20.8±1.2)%、(18.0±0.9)%和(0.2±0.1)%.结论 超声本身即有促进基因转染的作用,超声辐照联合微泡造影剂后则增强了这种作用,明显增加了基因的转染效率.     

叶酸靶向超声造影剂的制备及体外寻靶实验研究

目的 制备偶联叶酸的靶向超声微泡造影剂,鉴定其基本性质,并探讨体外寻靶能力.方法 将DSPE-PEG(2000)Folate溶入微泡成膜材料中,制备叶酸靶向超声微泡造影剂.用DFY型超声图像定量分析诊断仪检测微泡粒径和浓度.通过光镜和激光共聚焦显微仪观察该靶向造影剂对SKOV3细胞的体外寻靶情况,以普通超声微泡作对照.结果 靶向超声微泡的大小、粒径及分布与普通超声微泡相似.体外寻靶试验显示,该靶向微泡可以较多并牢固地聚集到SKOV3细胞表面,而普通微泡对照组未见特异性结合.结论 成功制备出偶联叶酸的靶向超声微泡造影剂.该造影剂在体外对高表达叶酸受体的SKOV3细胞具有较强的特异性亲合力,有望成为靶向显像卵巢癌的理想造影剂.     

自制脂质体超声纳米微泡的特性和显影效果

目的　测定自制纳米级脂质体超声微泡造影剂(nano-liposomal bubble,NB)的基本特性.方法　采用逆向蒸发法制备脂质体,用声震法制备NB后观察其形态,检测其平均粒径、表面电位及浓度等物理特性;在脱气水中分别注入生理盐水、NB及SonoVue (200μl),观察显影效果;分别经兔耳缘静脉团注生理盐水、NB及SonoVue (2.0 ml/kg),动态观察兔心脏及肝的增强情况.结果　镜下观察,NB粒径范围133.1～199.5 nm,平均粒径为(171.60±30.82)nm,平均电位为- (1.92±0.65)mV,分布均匀,浓度为(3.8～5.6)×108/ml.常温下造影剂放置1周、1月后观察,基本特性无明显改变.体外显影结果显示:NB与SonoVue一样具有显著的显影效果.体内造影后观察:NB与SonoVue均能明显增强兔心脏及肝的二维灰阶显像.结论　NB性质稳定,形态好,粒径均一,具有良好的显影效果.由于其粒径小且基于脂质体基础上易被修饰,因此在超声分子影像及基因/药物传递方面具有广阔的应用前景.     

微泡造影剂及超声促进绿色荧光蛋白质粒在人肝癌细胞HepG2内表达的实验研究

目的研究微泡造影剂与超声辐照是否能提高绿色荧光蛋白质粒在人肝癌细胞HepG2中的转染率。方法将培养的HepG2细胞分为四组：第一组为对照组；第二组以脂质体转染；第三组加入微泡造影剂SonoVue并予以超声辐照；第四组加入脂质体和微泡造影剂SonoVue并予以超声辐照．将合有绿色荧光蛋白报告基因的真核表达质粒pEGFP-N1转染人肝癌细胞HepG2，24小时后以荧光显微镜观察人肝癌细胞HepG2中的绿色荧光蛋白表达情况，并用流式细胞仪测算转染率。结果脂质体转染组与微泡造影剂＋超声辐照组有显著性差异（P〈0．05）。脂质体＋微泡造影剂＋超声辐照组与微泡造影剂＋超声辐照组有显著性差异（P〈0．01）结论微泡造影剂和超声辐照协同脂质体能提高目标基因在肝癌细胞内的转染率。     

靶向CD147纳米微泡的制备及超声成像能力检测

目的 制备靶向CD147纳米微泡造影剂,并评价其理化性质、对肝细胞肝癌(肝癌)细胞的靶向能力及超声成像能力.方法 将PLGA-PEG、DSPE-PEG2000-COOH、全氟丙烷饱和水溶液混合,通过偶联反应与CD147抗体结合,制备靶向CD147纳米微泡.将靶向CD147及非靶向纳米微泡分别与MHCC-97H(肝癌细胞...     

超声辐照微泡声学造影剂增强脂质体介导肝细胞基因转染的体外实验研究

目的研究超声破坏微泡声学造影剂提高脂质体介导肝细胞生长因子质粒（plRES—EGFP—HGF）在肝细胞中转染率的可行性。 方法将培养的肝细胞分为4组：（1）单纯对照组，（2）脂质体转染组，（3）超声辐照脂质体转染组，（4）超声辐照微泡＋脂质体转染组。第（4）组按照每孔加入造影剂剂量又分为1）10μl组，2）20μl组，3）30μl组，4）40μl组4亚组。超声辐照微泡＋脂质体转染组在脂质体介导肝细胞生长因子转染肝细胞1h后，在24孔板中每孔加入微泡声学造影剂10，20，30或40μl，超声辐照60s。24h后用荧光显微镜观察绿色荧光蛋白表达情况、MTT法检测肝细胞增殖率和流式细胞仪检测基因转染率。 结果超声频率1MHz，功率0．5W／cm。辐照60S，每孔加入造影剂30μl时肝细胞基因转染效率最高，与脂质体转染组比较有显著性。 结论在一定条件下，超声辐照微泡声学造影剂可明显提高脂质体介导肝细胞基因转染效率，为基因治疗提供了新的思路。     

含羧基末端连接臂脂质纳米微泡制备的探讨

目的制备纳米级含羧基末端磷脂微泡造影剂并筛选出最佳制备条件。方法采用薄膜水化-超声法通过改变超声声振时间、脂质成分比例以及聚乙二醇(PEG)的分子量、PEG与二硬脂酸-N-[羧基丙酰基(聚乙烯乙二醇-琥珀酰)磷脂酰乙醇胺](DSPE-PEG2000-COOH)的比例,通过观察超声造影剂的浓度、粒径及分布、稳定性,选出最佳制备方案。结果超声声振时间10 min、二棕榈酰乙醇胺(DPPE)与二棕榈酰磷脂酰胆碱(DSPC)质量比为3:1、DSPE-PEG2000-COOH与PEG4000比例为1:1,为最佳制备方案。结论制备含羧基连接臂的纳米脂质微泡,可为进一步制备靶向造影剂打下基础。     

人肝癌靶向脂质体微泡造影剂的鉴定及免疫学性质研究

目的 制备人肝细胞癌靶向免疫脂质体微泡造影剂，并对其免疫学性质进行研究。方法 静电吸附法制备免疫脂质体微泡造影剂；玻片凝集试验、荧光免疫染色试验证明抗体与脂质体微泡的结合；ELISA法检测免疫脂质体微泡的活性；花环形成试验、花环形成阻断试验及荧光免疫染色试验研究免疫脂质体微泡的靶向作用；细胞毒试验(MTT法)评价其安全性。结果 免疫脂质体微泡的玻片凝集试验、荧光免疫染色试验均为阳性；ELISA结果显示，免疫脂质体微泡中单抗的活性与游离单抗基本一致；免疫脂质体微泡围绕人肝癌细胞形成花环，花环形成率达90％以上；免疫脂质体微泡造影剂对人正常肝细胞的增殖没有影响。结论 采用静电吸附法成功制备了具有良好免疫活性的人肝细胞癌靶向免疫脂质体造影剂；该造影剂在体外能与人肝癌细胞高效特异结合，对人正常肝细胞没有毒性。     

低频超声联合微泡影响前列腺癌DU145细胞血管内皮细胞生长因子表达的实验研究

目的探讨低频超声联合微泡对前列腺癌DUl45细胞血管内皮细胞生长因子（VEGF）表达的影响。方法体外培养前列腺癌DUl45细胞,细胞呈对数生长时分为4组进行实验：对照组（细胞悬液1ml）、单纯微泡组（800μl细胞悬液加入200μl微泡剂SonoVue）、单纯低频超声组（1ml细胞悬液以频率80kHz、声强0．45W／cm2超声辐照30S）、低频超声联合微泡组（800μl细胞悬液加入200μl微泡剂SonoVue后立即以频率80kHz、声强0．45W／cm2超声辐照30s）。MTT法检测细胞存活率,流式细胞术检测凋亡率,Western印迹法检测细胞VEGF蛋白的表达。采用单因素方差分析比较对照组、单纯微泡组、单纯低频超声组、低频超声联合微泡组细胞存活率、凋亡率、VEGF蛋白表达量差异,进一步组间两两比较采用SNK-q检验。结果对照组、单纯微泡组、单纯低频超声组、低频超声联合微泡组细胞存活率分别为100％、（96．50±12．49）％、（93．20±5.31）％、（81．30±9．32）％;凋亡率分别为（1．10±0．26）％、（1．78±0．63）％、（5．85±0．45）％、（9．36±0．90）％;VEGF蛋白表达量分别为0．81±0．05、0．79±0．06、0．58±0．04、0．38±0．05。单纯微泡组、单纯低频超声组细胞存活率与对照组比较差异均无统计学意义,低频超声联合微泡组细胞存活率较对照组降低,且差异有统计学意义（q=5．88,P〈0．05）。单纯微泡组细胞凋亡率、VEGF蛋白表达量与对照组比较差异均无统计学意义;单纯低频超声组、低频超声联合微泡组细胞凋亡率均高于对照组,VEGF蛋白表达量均低于对照组,且差异均有统计学意义（q=14．09、24．51、8．06、14．89,P均〈0．05）。低频超声联合微泡组细胞凋亡率高于而VEGF蛋白表达量低于单纯低频超声组,且差异亦均有统计学意义（q=10．42、6．83,P均〈0．05）。结论微泡能增强低频超声抑制前列腺癌DUl45细胞增殖及促进凋亡,其机制可能与其下调前列腺癌DUl45细胞VEGF表达有关。     

六氟化硫脂质微泡的制备研究

目的 制备六氟化硫脂质微泡,为靶向微泡的研制奠定基础.方法 以二硬脂酰磷脂酰胆碱、聚乙二醇磷脂酰乙醇胺为膜成分,六氟化硫为气体核心,薄膜一超声法制备微泡.以SonoVue作对照,观测自制微泡的形态、粒径、浓度、pH值、渗透压等各项理化参数.在二次谐波模式下观察微泡显影体外水囊及兔肾实质,测量分析增强图像峰值灰度.结果 自制微泡和SonoVue均大小均匀,圆形,中空透亮,平均直径分别为2.25μm和2.50 μm,粒径分布分别为0.4～10 μm和0.2～10 μm,90%微泡直径分别控制在6 μm和8 μm以内.初始浓度分别为5 x 108～10 X 108/ml和1 x 108～5 x 108/ml,配置后的稳定性均为6 h.显影水囊灰度值分别为121.67±6.76和122.33±4.53(P>0.05),兔肾造影增强峰灰度分别为72.00±7.21和74.65±10.93(P>0.05).结论 脂质微泡制备成功,具有良好的理化特性及显影效果.     

PEGylated cationic liposomes robustly augment vaccine-induced immune responses: Role of lymphatic trafficking and biodistribution

Lymph nodes (LNs) are peripheral lymphoid organs essential for vaccine-induced immune responses. Although cationic liposomes have been documented as a novel adjuvant and vaccine delivery system, whether enhancing LN targeting would improve the efficiency of cationic liposome-formulated vaccines has not been elucidated yet. In the present study we investigated the effect of PEGylation on LN targeting and the immunogenicity of cationic liposome-formulated vaccines. DOTAP cationic liposomes were incorporated with 1 or 5mol% of DSPE-PEG2000 and labeled with near infrared fluorescent dyes. The lymphatic trafficking and biodistribution of different liposomes after subcutaneous (s.c.) injection were recorded using an in-vivo imaging system. The results showed that incorporation of 1mol% DSPE-PEG2000 not only accelerated the drainage of DOTAP liposomes into draining LNs, but also prolonged their LN retention and enhanced liposome uptake by resident antigen-presenting cells. On the other hand, although incorporating 5mol% of DSPE-PEG2000 into DOTAP liposomes enhanced their LN retention and uptake to a lesser extent, it prolonged blood circulation of DOTAP liposomes and increased their splenic accumulation. In addition, PEGylated DOTAP liposomes augmented primary and secondary anti-OVA antibody responses more potently than nonPEGylated DOTAP liposomes did. Hence, incorporating a small amount of DSPE-PEG2000 into DOTAP liposomes not only increased the passive LN targeting of DOTAP-formulated vaccines but also modulated their biodistribution in vivo, which consequently improved the efficiency of cationic liposome-formulated vaccines.     

Prednisolone retention in integrated liposomes by chemical approach and pharmaceutical approach.

The purpose of this study is to demonstrate a stable retention of prednisolone (PLS) in the unique liposomes integrated by lipophilic derivative approach and PEGylation approach. Palmitoyl prednisolone (Pal-PLS) was newly synthesized and used as a lipophilic derivative. The liposomes were composed of egg phosphatidylcholine (EggPC)/cholesterol (Chol) and L-alpha-distearoylphosphatidylcholine (DSPC)/Chol with or without L-alpha-distearoylphosphatidylethanolamine-polyethylene glycol 2000 (DSPE-PEG 2000) or -PEG 5000 (DSPE-PEG 5000). The retentions of PLS and Pal-PLS in the various liposomes were examined by ultrafiltration and gel filtration. Although PLS showed high trapping efficiency by all liposomes after ultrafiltration, low incorporation efficiency was observed in gel filtration. It indicates that PLS was released from the liposomes by a dilution with elution medium in gel filtration. Pal-PLS showed high incorporation into all liposomes after both ultrafiltration and gel filtration. The high incorporation of Pal-PLS into EggPC/Chol liposomes, however, was reduced by incubation with rat plasma in gel filtration. The reducing effect of rat plasma on drug incorporation into liposomes was inhibited by using DSPC and DSPE-PEGs. Thus, we systemically examined the drug retention in various liposomes and demonstrated the high retention of PLS in the liposomes integrated by lipophilic derivative approach and pharmaceutical approach using special lipids.     

低频超声照射SonoVue对绿色荧光蛋白质粒在小鼠肝癌移植瘤中表达的作用

目的 研究物理参数对体内基因转染率的影响,确定最佳剂量-效应关系.方法 建立小鼠(C57BL/6J)肝癌皮下移植瘤模型,经尾静脉注入绿色荧光蛋白质粒(pEGFP)和SonoVue混合物,给予不同辐照声强(1 W/cm2、2 W/cm2、3 W/cm2)、辐照时间(1 min、5 min、10 min)以及不同剂量SonoVue(30μl、60μl、90μl).流式细胞仪和荧光显微镜检测肿瘤细胞内绿色荧光蛋白表达.结果 pEGFP在肿瘤组织内的表达随辐照时间、辐照声强、造影剂剂量的增加而增多,持续增加辐照时间、声强和造影剂剂量并不能有效增加其转染率,2 W/cm2[(21.02±1.45)%]、5 min(23.22±1.91)%]、60μl SonoVue[(21.02±1.45)%]时,pEGFP在肿瘤组织内的表达达到最大.结论 不同物理参数对体内基因转染率有明显影响,2 W/cm2、5 min、60μl SonoVue的参数条件下,可达到最佳剂量-效应关系.     

绒毛组织冷冻切片与靶向超声微泡造影剂的结合特性研究

目的本研究旨在探讨耦合抗β-人绒毛膜促性腺素（β-HCG）抗体靶向超声微泡造影剂（TMCA）的稳定性及其与妊娠绒毛组织冷冻切片的结合特性。方法将制备好的TMCA与异硫氰酸荧光素（FITC）标记兔抗鼠IgG抗体稀释液混匀后,用流式细胞仪器检测不同时间点声诺维（SonoVue）微泡和抗β-HCG抗体的结合率。同时用妊娠绒毛组织冷冻切片、正常子宫内膜组织冷冻切片分别与TMCA、声诺维微泡进行结合反应,比较各组结合率、冲洗前后及抗β-HCG抗体预处理前后的结合率。结果流式细胞仪检测结果显示,声诺维微泡与FITC标记兔抗鼠IgG抗体的结合率随时间变化越来越高。寻靶实验发现,TMCA与绒毛组织、子宫内膜组织的结合率差异有显著的统计学意义[（83.30±2.15）%vs.（13.30±2.58）%,P〈0.01];绒毛组织与TMCA、声诺维微泡的结合率差异有显著的统计学意义[（83.30±2.15）%vs.（15.00±3.07）%,P〈0.01]。磷酸盐缓冲液（PBS）冲洗前后TMCA与绒毛组织的结合率差异无统计学意义[（83.30±2.15）%vs.（78.30±4.36）%,P〉0.05]。抗β-HCG抗体预处理前后TMCA与绒毛组织的结合率差异有显著的统计学意义[（83.3±2.15）%vs（16.7±2.5）%,P〈0.01）。结论 TMCA与绒毛组织在体外一定条件下能高效率、高强度、高特异性地稳定结合。     

Microbubble Stability is a Major Determinant of the Efficiency of Ultrasound and Microbubble Mediated in vivo Gene Transfer

In the search for an efficient nonviral gene therapy approach for the treatment of genetic disorders of cardiac and skeletal muscle such as Duchenne muscular dystrophy, ultrasound in combination with contrast enhancing microbubbles has emerged as a promising tool for safe and site-specific enhancement of gene delivery. Indeed, microbubble-enhanced gene transfer (MBGT) has been investigated for a wide variety of target sites using both reporter and therapeutic genes. Although a range of different microbubbles have been used for MBGT studies, comparison of their efficiencies is difficult because microbubble concentration and the ultrasound settings used for the application vary considerably. Only two studies to date have attempted a direct comparison of commercially available microbubbles, and both concluded that not all microbubbles show the same efficiencies with MBGT. Thus far, the reason for this is unclear. Here, the efficiency of three commercially available microbubbles—Optison, SonoVue and Sonazoid—was analyzed to understand the microbubble properties that are important for their function as an effective enhancer for gene transfer in vivo. In this study, plasmid DNA or antisense oligonucleotides were delivered by systemic injection with MBGT, focused on the heart. Gene delivery to the heart with equalized concentrations of the three microbubbles showed that Optison and Sonazoid are more efficient in MBGT compared with SonoVue, which showed the weakest gene transfer to the myocardium. Investigations into the properties of these microbubbles showed that size and shell composition did not directly influence MBGT, whereas the microbubbles with increased stability in an ultrasound field showed better MBGT results than those degrading faster. Moreover, the microbubble concentration used for MBGT was also found to be an important factor influencing the efficiency of MBGT. In conclusion, the stability of a microbubble was shown to be a major influential factor for its performance in MBGT, as is the concentration of the microbubbles used. These findings emphasize the importance of detailed investigations into the properties of microbubbles to allow the production of a microbubble specifically designed for optimum performance with MBGT. (E-mail: d.wells@imperial.ac.uk)     

超声微泡连接特异性抗体的制备方法及靶向化处理因素对超声微泡物理性质的影响

目的 探讨超声微泡连接特异性抗体的制备方法及靶向化处理因素对超声微泡物理性质的影响.方法 应用生物素-链霉亲和素连接系统,使注射用六氟化硫微泡(SonoVue)与特异性抗血管细胞黏附分子-1(VCAM-1)抗体相连接,用间接免疫荧光法检测抗体与微泡的连接,用马尔文激光粒径分析仪分别测定生物素化处理前后微泡的粒径,采用超声诊断仪评价微泡靶向化构建前后的显像效果.结果 SonoVue与特异性抗体成功连接,间接免疫荧光检测呈阳性.生物素修饰后的微泡粒径分布变窄,与普通微泡的超声显像效果略有差异.结论 通过生物素-链霉亲和素系统可以成功靶向化构建超声微泡,靶向化处理因素对微泡的粒径有一定影响,对微泡的超声显影效果略有影响.     

超声造影剂SonoVue介导质粒绿色荧光蛋白转染小鼠骨骼肌细胞的实验研究

目的 探讨微泡造影剂SonoVue介导基因转染小鼠骨骼肌细胞的作用。方法 采用质粒绿色荧光蛋白（GFP）作为目的基因，超声结合SonoVue作用于小鼠H2K成肌细胞，照射时间分别为10s、20s、30s、40s、50s、60s，流式细胞仪测定GFP阳性细胞率，台盼蓝染色测定细胞生存率。SonoVue结合或不结合超声作用于小鼠胫前肌，1周后处死小鼠，荧光显微镜测定GFP阳性肌纤维数，HE染色估计肌肉破坏面积。结果 加入SonoVue后，GFP阳性细胞率与细胞生存率总体显著低于阳性对照组；动物实验显示，SonoVue结合或不结合超声均显著增强GFP基因表达水平作用。结论 体外细胞培养SonoVue无增强GFP基因转染的作用，却增加细胞损伤；活体小鼠实验显示SonoVue具有良好的增强骨骼肌细胞基因表达作用。     

Evaluation of circulation profiles of liposomes coated with hydrophilic polymers having different molecular weights in rats.

The purpose of this study was to evaluate the circulating properties of liposomes coated with modified polyvinyl alcohol (PVA-R) having different molecular weights (6000, 9000 and 20000). The size controlled liposomes (egg phosphatidylcholine (or distearoylphosphatidylcholine):cholesterol=7:3 in a molar ratio) were prepared by the hydration method followed by sonication. Polymer coated liposomes were prepared by just mixing the resultant liposomal suspension and a polymer solution. The effects of polymer coating were evaluated by measuring the circulation time of the injected liposomes after i.v. administration in rats and the dispersing property of the liposomes in a biological condition. The circulation of the PVA-R coated liposomes was prolonged with increasing the molecular weight of PVA-R. The aggregation and/or fusion of the liposomes in the presence of serum in vitro was also depressed more by coating the liposomes with PVA-R having higher molecular weight. There was a good correlation between the circulation time and the physical stability of non-coated and the various PVA-R coated liposomes. The prolonged circulation time of PVA-R (molecular weight: 20000) coated liposomes (ca. 1.3 mol% coating) was comparable to that of a stealth liposome prepared with 8 mol% of DSPE-PEG (molecular weight of PEG: 2000).