Obtain a rapid burst of Cre expression in mammalian cells with footprint-free delivery of Cre mRNA in nonintegrating, virus-like particles. Cre RNA lentiviral particles (RLPs) contain no viral genome and are generated using a novel packaging technology that loads multiple biologically active mRNAs per particle. RLPs have been shown to mediate transient protein expression at high efficiency for in vitro and in vivo applications involving cell lines, primary cells, stem cells, tissues, and organisms.
Obtain a rapid burst of Cre expression in mammalian cells with footprint-free delivery of Cre mRNA in nonintegrating, virus-like particles. Cre RNA lentiviral particles (RLPs) contain no viral genome and are generated using a novel packaging technology that loads multiple biologically active mRNAs per particle. RLPs have been shown to mediate transient protein expression at high efficiency for in vitro and in vivo applications involving cell lines, primary cells, stem cells, tissues, and organisms.
Popular methods for Cre recombinase-based genome editing present tradeoffs. Transient transfection of a plasmid or mRNA limits the scope of Cre activity, minimizing the likelihood of unwanted effects associated with persistent Cre expression, but is associated with cellular toxicity, is unsuitable for in vivo applications, and is only feasible for cells that can be readily transfected. Transduction of the Cre gene sequence on a viral vector enables Cre expression in a broader range of cell types and can be applied to in vivo studies, but is associated with random genomic integration, may require activation and/or selection of target cells, and may result in undesirable persistence of Cre expression.
Cre recombinase RLPs enable researchers to overcome the above limitations and express Cre more rapidly and efficiently in mammalian cells by delivering nonviral Cre mRNA in lentivirus-derived packaging in the absence of a viral genome. In vitro studies involving RLP-mediated delivery of a luciferase reporter demonstrated that reporter expression was maximal at 4–24 hours post application and had declined rapidly at 30 hours. By contrast, expression of luciferase delivered via transfection was undetectable until 8 hours and peaked at 24 hours, while expression via lentiviral integration was undetectable until 24 hours. In an in vivo study involving a loxP mouse line (ROSA26-YFP), local injection of Cre RLPs into muscle yielded a zone of reporter gene expression that was comparable in size to a zone generated via injection of purified Cre lentivirus (Prel et al. 2015).
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其次，察看次目的蛋白的存在形式，有没有多聚体形式及变构形式；
最后，查看多家抗体公司的DATA，看看别人的WB做出来的条带的位置。
根据你说的，特异识别多个组织中的同样大小的条带，我觉得很可能就是你的目的蛋白。
一般来说多克隆的阳性率高一些，但出现假阳性的比例也高一些。
1取组织
（1） 灌注固定：暴露心脏,将注射针头插入左心室,剪开右心耳,快速灌注PBS,待右心耳处流出澄清液体,换4%多聚甲醛灌注到大鼠四肢强直为止,快速取脑和肾。
（2） 固定：将组织置于4%多聚甲醛中固定15小时
（3） 脱水：将组织放入30% 蔗糖溶液中直至下沉
（4） 切片：脑组织冰冻切片20微米，37度烘干2h以上；
（5）将切片放置－80度保存，随用随取。
2免疫单标记
（1）做抗原修复（0.4g柠檬酸、3g柠檬酸钠、1000ml水配制成溶液，放到水浴箱中90~94℃保存15min）,冷却后， 用1*PBS反复洗标本10min，吸去PBS，同时向标本加渗透液（配0.4%trition液，然后将BSA加入到配好的trition液中，配成1％浓度）100ul，常温下湿盒反应1h。
（2） 吸去渗透液，同时向标本加block液（5％封闭用正常山羊血清）100ul，常温下湿盒反应5h。
（3） 吸去block液，同时向标本加入1：50稀释的羊抗鼠一抗100ul，37度湿盒反应过夜。
（4）吸去一抗，并回收，用1*PBS洗液反复洗15min。
（5）快速从冰箱中取出驴抗羊的二抗，并立即用锡纸包裹，注入到一个新的以锡纸包裹的tube管中并用PBS以1：200稀释。
（6）进入暗室，吸去PBS，同时加入二抗100ul，37度湿盒反应2h,封片。
（7）放置到荧光显微镜下观察。
又由于自然存在的抗原大都存在多个抗原表位,会刺激机体产生多种针对同一抗原的不同抗原表位相应的不同抗体.
