
Description
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Transfection Reagent for SK-MEL-28 Cells (Melanoma Cells, HTB-72)
- A nanoparticle-based liposome formulation
- Transfection protocols provided for transfection of proteins, DNA, mRNA, siRNA, shRNA and microRNA
- Transfection Enhancer reagent provided with the kit
- Produce higher level of recombinant protein expression with minimal disruption of normal cell function
- Generate physiologically relevant data you can trust
- Easy-to-use transfection protocol with reproducible results
- Low cytotoxicity
- Download in vitro SK-MEL-28 transfection protocol: [PDF]
- Download SK-MEL-28 CRISPR/Cas9 transfection protocol: [PDF]
- Download PowerPoint presentation for SK-MEL-28 cells transfection kit: [PPT]
- Developed and manufactured by Altogen Biosystems
Transfection Efficiency:
Reagent exhibits at least 71% transfection efficiency of siRNA delivery. Transfection efficiency was determined by qRT-PCR.
Transfection Protocol and MSDS:
Download Altogen Biosystems SK-MEL-28 Transfection Protocol: [PDF]
Download MSDS: [PDF]
SK-MEL-28 Cell Line:
Malignant melanoma is an invasive form of skin cancer that originates from the transformation of melanocytes. It is characterized by the highly aggressive nature and ability to metastasize to diverse distant sites. Human cell lines can provide a realistic prediction of drug responses in humans as well as information on safety and efficacy of drug candidates. The SK-MEL-28 cell line was established from the skin of a 51-year-old male patient with malignant melanoma. SK-Mel-28 cells exhibit polygonal cell morphology, and this is just one of many melanoma cell lines (also named SK-MEL). The modal chromosome number of 90 occurs nearly in 50% of cells. Also, the hypotetraploid SK-MEL-28 expresses wildtype N-Ras gene. This tumorigenic cell line could be helpful for skin cancer research as well as for other cell and molecular biology applications. Altogen Biosystems manufactures nanoparticle-based transfection kits for the SK-MEL-28 cell line.
Data:
Figure 1. siRNAs targeting Lamin A/C mRNA or non-silencing control siRNA were transfected into SKMEL28 cells following the recommended protocol. At 48 hours post-transfection the cells were analyzed by qRT-PCR for Lamin A/C gene expression levels. 18S rRNA levels were used to normalize the Lamin A/C data. Values are normalized to untreated sample. Data are means ± SD (n=4).
Figure 2. Protein expression of Lamin A/C in SK-MEL-28 cells. DNA plasmid expressing Lamin A/C or siRNA targeting Lamin A/C were transfected into SK-MEL-28 cells following Altogen Biosystems transfection protocol. At 72 hours post-transfection the cells were analyzed by Western Blot for protein expression levels (normalized by total protein, 10 µg of total protein loaded per each well). Untreated cells used as a negative control.
Selected in vivo transfection product citations (ALTOGEN® IN VIVO Transfection Kits used in the following publications):
- 2008 454(7203):523-7. Innate immunity induced by composition-dependent RIG-I …Saito et al [PDF]
- Am J Pathology. 2010 177(4):1870-80. Role of ocular complement factor H in a murine model … Lyzogubov et al [PDF]
- Nature Biotechnology. 2011 29(4):341-5. Delivery of siRNA to the mouse brain by … Alvarez-Erviti et al [PDF]
- Cancer Research. 2011 71(15):5144-53. Inhibition of miR-193a expression by… Iliopoulos et al [PDF]
- 2010 16(11):2108-19. RNase L releases a small RNA from HCV RNA that refolds … Malathi et al [PDF]
- 2012 55(7):2069-79. The p47phox- and NADPH oxidase organiser 1 … Youn et al [PDF]
- British Journal of Cancer. 2012 107(3):516-26. TIGAR induces p53-mediated cell-cycle … Madan et al [PDF]
- 2014 63(2):353-61. Tissue transglutaminase contributes to … Liu et al [PDF]
- Circulation Research. 2010 15;107(8). Kruppel-like factor-4 transcriptionally regulates … Cowan et al [PDF]
- 2012 59(1):158-66. Role of uncoupled endothelial nitric oxide synthase … Gao et al [PDF]
- Jounal of Biological Chemistry. 2012 287(4):2907. Chaperoning of mutant p53 protein … Gogna et al [PDF]
- PLoS Pathogens. 2012 8(8) Uridine composition of the poly-U/UC tract of HCV RNA … Schnell et al [PDF]
- J Proteome Res. 2012(11) Retinal proteome analysis in a mouse model of oxygen-induced … Kim et al [PDF]
- 2011 141(2) Differential type I interferon-mediated autophagic trafficking … Desai et al [PDF]
- PLoS Pathog. 2014 10(10) Exosomes from hepatitis C infected patients transmit HCV … Bukong et al [PDF]
SK-MEL-28 Transfection Kit
Altogen Biosystems:
Altogen Biosystems provides pre-optimized transfection kits and electroporation products for life science research. Transfection products are developed for individual cancer cell line and transfection protocols are optimized to enable high transfection efficiency of biomolecules. Altogen Biosystems developed in vivo delivery products for small animal research, mouse and rat targeted tissue delivery: liver targeted, pancreas targeted, kidney targeted, PEG-Liposome-, Nanoparticle-, Lipid-, and Polymer-based in vivo transfection kits. Advanced formulation of reagents and optimized transfection protocols provide efficient intracellular delivery of biomolecules. Read more about transfection technology at Altogen’s Transfection Resource.
Altogen Labs Research Services:
Altogen Labs provides GLP-compliant contract research studies for pre-clinical research, IND applications, and drug development. Biology CRO services include: Xenograft models (30+), development of stable cell lines, ELISA assay development, cell-based and tissue targeted RNAi studies, safety pharm/tox assays, and other studies (visit AltogenLabs.com).
Volume Options:
- 0.5 ml (Catalog #6908)
- 1.5 ml (Catalog #6909)
- 1.5 ml CRISPR (Catalog #2193)
- 8.0 ml (Catalog #7086)
AltogenBiosystems是一家开发和制造用于生命科学研究,药物发现和开发的转染试剂盒的生物技术公司。转染试剂盒针对特定癌细胞系和原代细胞培养进行了优化,可将生物分子有效递送到靶组织中。通过先进的试剂配方和优化的转染方案实现体外(癌细胞系)和体内(动物组织靶向试剂、癌细胞系)递送货物分子,包括质粒DNA,各种类型的RNA(mRNA,siRNA,shRNA,microRNA),蛋白质和小分子研究。
Altogen生命科学公司致力于研发,生产和销售特定细胞系的转染试剂,用于细胞间生物分子的传递,并通过对转染试剂类型的设计将siRNA和质粒DNA有效地转入不同的细胞系和原代细胞内。Altogen公司开发的聚合物,脂质体,纳米粒子为基础的转染技术分别针对分子生物学,组合化学,和细胞生物学而分别应用。Altogen定制服务提供符合GLP要求定制研究服务,包括代稳定的细胞系,细胞银行和冷冻保存,焦磷酸测序,克隆,RNA干扰(RNAi)和基因沉默服务,发展分析,siRNA文库筛选,并转染服务。稳定的肿瘤细胞株和原代细胞的产生,可以是非常昂贵和费时。该公司的细胞培养科学家的细胞株的选择,无论是利息或shRNA表达载体的稳定表达的基因改造。标准的RNAi技术服务,包括设计与合成的siRNA的利益,验证siRNA的沉默效率,siRNA转染条件的优化,使高效的基因沉默细胞系或原代培养细胞的靶基因。转染培养细胞的瞬时或稳定的引入外源性分子和遗传物质(即RNA或DNA),通常是在生物实验室用来研究基因功能,基因表达的调节,生化映射,突变分析,和蛋白质的生产。科学家利用各种载体分子,这种分子,使质粒DNA(PDNA),信使RNA(mRNA),短干扰RNA(siRNA),小分子RNA(miRNA)的,并进入肿瘤细胞株和原代细胞的蛋白质的基因交付。不幸的是,无单提货的方法或转染试剂,可以适用于所有类型的细胞,细胞的细胞毒性和转染效率显着不同,取决于试剂,协议,并正在利用细胞类型。Altogen生物系统公司提供超过60种类型的细胞的预优化转染试剂盒。纳米粒子,脂质和聚合物基ALTOGEN®在体内转染试剂,使交付功能的RNA和DNA分子在体内。PEG脂质体在体内输送系统减少由于PEG修饰的先天免疫反应,并提供高效的siRNA转染的DNA,并在体内的蛋白质。由科学“杂志(2010年12月17日):PEG脂质体在体内转染试剂盒siRNA的特色Altogen生物系统功能的特定细胞系转染试剂盒
120+细胞转染试剂和活体组织靶向试剂盒制造商AltogenBiosystems是一家生物技术公司,开发和制造用于生命科学研究、药物发现和开发的转染试剂盒。Altogen®体内转染试剂可有效地将生物分子导入靶组织。细胞转染试剂盒针对特定的癌细胞系和原代细胞进行了优化。通过先进的试剂配方和优化的转染方案实现货物分子(DNA、RNA、蛋白质)的高效传递。AltogenBiosystems利用高分子化学、分子和细胞生物学的专业知识,开发了新的体内外给药技术。转染是将外源分子导入培养细胞中,常用于研究基因功能、基因表达调控、生化定位和蛋白质生产。不幸的是,由于细胞毒性和转染效率的差异很大,并且取决于所使用的试剂、方案和细胞类型,因此没有一种单一的传递方法或转染试剂可应用于所有类型的细胞。AltogenBiosystems为120多个癌细胞系和原代细胞类型提供优化的转染试剂盒和电穿孔产品。体内转染试剂可实现组织靶向给药。Altogen的转染试剂盒包括用于体外(癌细胞系)和体内(用于动物研究的组织靶向试剂)转染的转染增强剂试剂和转染复合物冷凝器。Altogen实验室提供符合GLP的实验室合同研究服务。我们的生物CRO服务包括异种移植物的疗效、IND应用的pharm/tox研究和安全性测试、分析开发(ELISA、IC-50、qPCR)、90多个异种移植物动物模型、RNAi和基因沉默服务。Altogen的细胞培养科学家通过在28天内培育出稳定的细胞系,将选择的细胞系转化为稳定表达感兴趣的基因。
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的抗体不容易染色,哪为同仁有这方面的经验?另外基因公司卖的cellsignalingtechnology的抗体要1875元,而中山公司卖的这个抗体只有800多,其中差距在哪里?请赐教
GlucosestarvationcausestranslocationofAMPKβ2tothelysosomeinHEK-293cellsthatisdependentonN-myristoylation.Theexperimentwasperformedinβ2KOcellsasinFig.1c,exceptthatthelysosomalMarkerLAMP1(taggedwithRFP)wasco-expressedwiththewild-typeormutantAMPKβ2.Upperpanelsshowmergedimagesstainedblue(4′,6-diamidino-2-phenylindole(DAPI),nuclei),red(LAMP1,lysosomes)andgreen(AMPKβ2,detectedusingantibodyvalidatedine),incellsincubatedwithorwithoutglucosefor20 min.Lowersmallpanelsaremagnificationsoftheareasindicatedbydashedboxesintheupperpanels,showing(LtoR)redandgreenchannelsandmergedimages.
下面的这段话是图注,图注的意思我明白,但是我想知道merge后的图看什么颜色的荧光,蓝色是细胞核,红色是lysosome(位于胞质),绿色是AMPKβ2,该实验是想观察AMPKβ2是否转位到lysosome上了,如果确实发生了AMPKβ2转位到lysosome上,那么merge后是红色与绿色融合在一起,是吗?融合在一起发什么颜色的光了?
1)做免疫组化的片子最好不要同时做HE染色,因为那样会掩盖阳性着色的结果;而且用苏木素复染细胞核也是淡染,若要在免疫组化的片子上观察形态学的改变,除非有非常典型的表现,一般是有很大的难度的。
2)你是培养的细胞做免疫组化吗?如果是的话,那就每个样本只有一张片子吧?唯一的办法是进行免疫双染,同时做CD34和AFP(甲胎蛋白)。我觉得后者阳性便可以或多或少的获得向肝细胞分化的证据。
3)用HE染色来识别造血干细胞向肝细胞分化,也就是说要看到肝细胞的典型形态学改变后才可以下结论。问题是:早期的肝细胞仅凭形细胞态学观察并不能很容易的被识别。病理大夫看组织切片,重要的是看其组织学结构,对肝细胞也是这样。如果单拿出来一个细胞,还是培养的细胞,染色后真的不容易区分。所以我觉得你设计的用HE染色来确定早期细胞向哪个方向分化不是很有说服力。
4)如果是切片,也就是说每个样本可以有多张切片,那可以在连续切片上分别染CD34和AFP,拍照时选择相同视野,也有一定说服力。但是还是不如双染法更可靠。
5)用双染最大的问题在于:这两者都表达在胞浆中,染色容易相互覆盖。所以在双染的方法选择上你还要再考虑一下,可不可以用免疫荧光双染?这样也很有说服力的
免疫组织化学又称免疫细胞化学,是指带显色剂标记的特异性抗体在组织细胞原位通过抗原抗体反应和组织化学的呈色反应,对相应抗原进行定性、定位、定量测定的一项新技术。它把免疫反应的特异性、组织化学的可见性巧妙地结合起来,借助显微镜的现像和放大作用,在细胞,亚细胞水平检测各种抗原物质,并可在原位显示相应的基因和基因表达产物。免疫组织化学技术现已有:免疫荧光组织(细胞)化学技术、免疫酶组织(细胞)化学技术、亲和组织化学技术、免疫金银及铁标记免疫组织化学技术等。免疫荧光组织(细胞)化学技术是采用荧光素标记的已知抗体(或抗原)作为探针,检测待测组织、细胞标本中的靶抗原(或抗体),形成的抗原抗体复合物上带有荧光素,在荧光显微镜下,由于受高压汞灯光源的紫外光照射,荧光素发出明亮的荧光,这样就可以分辨出抗原(或抗体)的所在位置及其性质,并可利用荧光定量技术计算抗原的含量。以达到对抗原物质定位、定性、定量测定的目的。

