抗EGFR抗体[ ep38y ](ab52894)

种属反应性

与反应:小鼠、大鼠、人
  • 应用 AB评论 说明 WB 1 / 1000 – 1 / 10000。检测到的频带的约175 kDa(预测分子量:134 kDa)可阻断。表皮生长因子受体的肽(ab204282)

    本产品采用A431了Western blot的强烈信号(鳞癌)裂解液的自然表达EGFR蛋白。Western blot检测条件可能需要优化的细胞和组织中表达较低水平的内源性表皮生长因子受体

    IP 1 / 50。 流式细胞 1 / 100。

    ab172730兔单克隆抗体,适用于该抗体的同型对照。

    ihc-p 用于测定浓度依赖性。

    小鼠和大鼠的推荐是基于WB结果。这种抗体可能不适合应用免疫组化方法检测小鼠或大鼠样本。

    国际商会/如果 1 / 250 – 1 / 500。
  • 靶标

    • 功能酪氨酸激酶受体结合的配体的EGF的家人和几个信号转导通路的激活细胞外信号转换成适当的细胞反应。已知的配体包括EGF、TGFa/TGFα,双调蛋白,纯化/苏州,BTC /β细胞素,表皮调节素/ EREG和HBEGF /肝素结合表皮生长因子。配体结合导致受体同源和/或异二聚体和关键残基磷酸化胞浆。磷酸化的受体蛋白Grb2新兵适配器从而激活下游的信号转导通路的复杂。激活至少4种主要的下游信号通路包括RAS-RAF-MEK-ERK,PI3激酶Akt,plcgamma PKC和统计模块。也可以激活序列的信号级联。也直接磷酸化蛋白质像RGS16,激活其GTP酶的活性可能是耦合的表皮生长因子受体信号转导的G蛋白偶联受体信号。也会增加其与MUC1和Src和CTNNB1 /β-连环蛋白。2型可以作为EGF作用的拮抗剂。
    • 组织特异性广泛表达。2亚型也在卵巢癌中表达。
    • 疾病相关肺癌皮肤和肠道炎性疾病、新生儿、2
    • 序列相似性属于蛋白激酶家族。酪氨酸蛋白激酶家族。表皮生长因子受体家族。包含1个蛋白激酶结构域。
    • 翻译后修饰在ser-695磷酸化是局部的,只有thr-693磷酸化发生。在thr-678和thr-693 PPKD1磷酸化的抑制EGF诱导MAPK8 / JNK1的激活。ptprj防止去磷酸化的内吞作用和稳定在细胞膜受体。自身磷酸化在tyr-1197被甲基化在arg-1199刺激和增强互动PTPN6。在tyr-1092和/或tyr-1110新兵STAT3磷酸化。去磷酸化和PTPN2 PTPN1。单泛素化和多聚泛素化的表皮生长因子刺激后;不影响酪氨酸激酶活性或信令能力但可能在溶酶体中发挥作用。多聚泛素连接主
      要是通过“赖氨酸-63”,而是通过“lys-48联动”、“lys-11 '和' lys-29还产生。
      去泛素化降解otud7b防止。泛素化的rnf115和rnf126。甲基化。甲基化的arg-1199 PRMT5刺激磷酸化在tyr-1197。
    • 细胞定位分泌和细胞膜。内质网的膜。高尔基体膜。核膜。内体。内体膜。核。针对表皮生长因子,进入细胞膜,高尔基体和内质网的核通过。内吞后激活的配体。共存与雌激素激动剂诱导的癌相关成纤维细胞的核gper1(CAF)。
    • 以上信息来自:UniProt加入目标p00533UniProt协会通用蛋白质资源(UniProt)2010
      核酸研究38(2010):d142-d148

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    日本进口动物安全标记笔Animal Marker

    日本进口动物安全标记笔Animal Marker

    动物安全标记笔Animal Marker





    使用日本厚生省法定色素,无毒性,经急性经口实验验证对实验动物无伤害

    动物安全标记笔Animal Marker :

    CatNo. 蓝 绿 红 黄 橙
    容量  AM15:15 ml
    包装数量 1盒:10支装 (各2支 / 色  可自由配色)

    动物安全标记笔:

    1,可简便并放心使用的标记笔,使用日本厚生省法定色素。

    2,配备了5种高识别度色彩(蓝,绿,红,黄,橙)。

    3,对动物皮肤、体毛的渗透力较强,即便被舔舐也不易掉色。

    4,一支标记笔用于实验动物老鼠上大致可处理400-600只。

    5,绝对保证长期试验过程中不褪色,在一般条件下可保持6-12周的良好辩识度。

     

    上海金畔生物科技有限公司提供5202一次性小鼠灌胃针 M340444

    日本进口小鼠软管一次性灌胃针

    5202一次性小鼠灌胃针 M340444 详情

    日本进口Ptfe软管材质 一次性小鼠灌胃针

    货号 规格 针长 针头 外径*内径 针基色 销售单位
    5200S 小鼠用 30 1.9 0.90*0.50 250支(5支/小包)箱
    5200 小鼠用 38 1.9 0.90*0.50
    5202SS 小鼠用 25 2.0 1.18*0.68
    5202S 小鼠用 30 2.0 1.18*0.68
    5202 小鼠用 38 2.0 1.18*0.68
    5204 大鼠用 52 2.8 1.79*1.19
    5206 大鼠用 78 2.8 1.79*1.19

     

    此灌胃针使用Ptfe软管材质,前端附有硅胶软球。可以有效减少对肺部以及邻近器官的损伤。操作简便。

    适用于多种小型实验动物,规格齐全,更可根据您的需求尺寸进行定制。

    5根配套小包装,EOG灭菌。

     

    固话总机:021-50837765
    订货热线:15221999938

    qq号:2743691513 1042640511

    微信号:jinpanbio
    网 址: www.jinpanbio.com
    金畔博客:www.jinpanbio.cn

    Email:sales@jinpanbio.com

    更多 www.jinpanbio.com.cn  www.utopbio.com

    日本药典,考试用试剂|医药/化妆品|关东化学株式会社

    日本药典依据试剂,考试用的试剂,关于生产销售。

    弹出窗口打开

    3局对应特级试剂

    日本药典,考试用试剂|医药/化妆品|关东化学株式会社

    医薬品考试全球能对应的产品,所以广泛活用吗。

    制剂稳定性试验用试剂

    日本药典,考试用试剂|医药/化妆品|关东化学株式会社

    崩溃考试及溶出考试能使用的试剂准备着。

    日局对应标准液

    日本药典,考试用试剂|医药/化妆品|关东化学株式会社

    日本药典的处方,按照规定的浓度的制备的各种标准液为您准备。

    硫酸呈色物试验用试剂

    日本药典,考试用试剂|医药/化妆品|关东化学株式会社

    硫酸呈色物考试是日本药典中医药品中的微量杂质检查有效的分析方法之一被采用。

    残留溶剂试验用试剂

    日本药典,考试用试剂|医药/化妆品|关东化学株式会社

    甲醇等低沸点不纯物含量降低溶剂,残留适合性试验,质量保证。

    液小黑溶剂

    3局对应HPLC纯试剂

    日本药典,考试用试剂|医药/化妆品|关东化学株式会社

    医薬品考试全球能对应的产品,所以广泛活用吗。

    医药/化妆品的咨询

    信息查询

    电话: 03 – 6214 – 1093

    传真: 03 – 3241 – 1054

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    MultigeneTM optimax PCR仪

    品牌:美国Labnet

    订货号:TC9610-230V

    产品介绍

    Labnet新款MultiGene Optimax PCR仪带来了新的运行速度和功能。该款机器继承了MultiGene原先简单的操作界面,并拥有更快的升降温速度,大大减少反应过程的运行时间。增加了PCR viewerTM计算器等功能。PCR viewer通过USB接口与仪器连接使用,透过PCR viewer窗口,可以实时查看实际的温度变化曲线。全新采用了6 – 4×4peltier排列格式,将整个反应模块划分成6个独立的区域,可以在每方阵设置独立的温度进行最佳条件摸索,或者进行不同温度的PCR反应。你不须选择高温和低温,而是让系统完全按照你的要求设置,为研究提供更大的灵活性。 Labnet  MultiGene Optimax PCR仪,以实惠的价格给您带来卓越的性能。

     

    二、特点

         快速升降温速度。

         热盖压力恒定,滑盖防止烫伤;可立便于清洁。

         热盖2种温度范围可选。

         全新热盖设计,高度防止冷凝水设计。

         6个独立控制的Peltier 模块。

    • 上海金畔生物科技有限公司

      文章号:441-441

    PL-024 食品级1.5mm物料PU软管 – 食品、医药行业应用

    PL-024 食品级1.5mm物料PU软管 – 食品、医药行业应用

    颜色

    透明,带有PU覆盖层的螺旋状镀锌钢丝加强

    温度

    -40°C/+ 90°C(短时间可耐受125°C)

    特性

    聚醚型聚氨酯软管, 带有PU覆盖层的螺旋状镀锌钢丝加强筋。内壁平滑以保证物料在管内流动顺畅。壁厚均为1.5毫米。耐霉菌、耐水解。具有优异的耐臭氧和耐候性能。符合FDA, EU10/2011。该软管无毒、无味、不含卤素,符合RoHS环保指令的要求。螺旋钢丝接地可以排静电,符合TRB S 2153的规定。阻燃度符合UL 94 HB标准。

    应用

    食品加工行业、制药行业、割草机和扫地清洁车等需要耐霉菌和耐水解的应用。可用于油脂类食品的输送。

    • 产品详情
    产品编码 内径 壁厚 耐负压 弯曲半径 体积 重量 包装长度
    No. in. mm mm mH2O mm m3 kg/m m
    PL-024-1000 1″ 25 1.5 9.5 30 0.041 0.305 20
    PL-024-1250 1-1/4″ 32 1.5 9.5 35 0.056 0.435 20
    PL-024-1500 1-1/2″ 38 1.5 9.5 40 0.074 0.465 20
    PL-024-1580 1-9/16″ 40 1.5 9.5 50 0.077 0.485 20
    PL-024-1770 * 45 1.5 9.0 50 0.090 0.545 20
    PL-024-1970 * 50 1.5 9.0 60 0.099 0.600 20
    PL-024-2000 2″ 51 1.5 9.0 60 0.106 0.610 20
    PL-024-2360 2-3/8″ 60 1.5 8.5 70 0.122 0.710 20
    PL-024-2500 2-1/2″ 63 1.5 8.0 70 0.148 0.745 20
    PL-024-2560 2-9/16″ 65 1.5 7.5 70 0.152 0.765 20
    PL-024-2760 * 70 1.5 7.5 80 0.165 0.920 20
    PL-024-2950 * 75 1.5 7.0 80 0.176 0.985 20
    PL-024-3000 3″ 76 1.5 7.0 90 0.232 0.995 20
    PL-024-3150 * 80 1.5 6.0 90 0.254 1.045 20
    PL-024-3540 * 90 1.5 5.0 100 0.294 1.170 20
    PL-024-3940 * 100 1.5 4.0 120 0.281 1.295 15
    PL-024-4000 4″ 102 1.5 3.5 120 0.286 1.320 15
    PL-024-4330 * 110 1.5 3.2 130 0.307 1.420 15
    PL-024-4720 * 120 1.5 3.0 140 0.333 1.545 15
    PL-024-5000 5″ 127 1.5 2.9 150 0.405 2.015 15
    PL-024-5120 * 130 1.5 2.6 150 0.414 2.065 15
    PL-024-5510 * 140 1.5 2.4 160 0.504 2.215 15
    PL-024-5910 * 150 1.5 2.0 170 0.537 2.370 15
    PL-024-6000 6″ 152 1.5 2.0 170 0.544 2.400 15
    PL-024-6300 * 160 1.5 1.8 180 0.662 2.525 15
    PL-024-7090 * 180 1.5 1.5 210 0.855 2.580 15
    PL-024-7870 * 200 1.5 1.2 230 1.130 2.860 15
    PL-024-8000 8″ 203 1.5 1.2 230 1.146 2.900 15
    PL-024-8980 * 228 1.5 1.0 260 0.933 3.250 10
    PL-024-9840 * 250 1.5 0.8 290 1.170 3.555 10
    PL-024-10000 10″ 254 1.5 0.8 290 1.188 3.610 10
    PL-024-10430 * 265 1.5 0.7 300 1.238 3.765 10
    PL-024-11000 11″ 279 1.5 0.7 320 1.301 3.960 10
    PL-024-12000 12″ 305 1.5 0.7 350 1.618 5.245 10

    产品名称:PL-024 食品级1.5mm物料PU软管
    行业应用:PL-024 食品级1.5mm物料PU软管 应用透明,带有PU覆盖层的螺旋状镀锌钢丝加强

    尘埃落定!贝瑞和康借壳上市获证监会无条件通过

    尘埃落定!贝瑞和康借壳上市获证监会无条件通过

    中国证券监督管理委员会(证监会)上市公司并购重组审核委员会在今日上午召开的第19次会议中,审核通过了成都天兴仪表股份有限公司发行股份购买资产的相关事宜,标志着基因测序领先企业贝瑞和康借壳上市取得了实质性的成功。

    尘埃落定!贝瑞和康借壳上市获证监会无条件通过

    下面我们来系统回顾本次并购重组过程中的几个重要阶段:

    2016年6月

    成都天兴仪表股份有限公司发布停牌公告,透漏公司将筹划重大资产重组事项,至此,开始长达6个月的停牌过程。

    2016年8月

    天兴仪表的重大资产方案首次披露:天兴仪表将其目前拥有的全部资产、负债、业务、人员等出售给天兴集团或其指定的第三方,天兴集团或其指定的第三方以现金形式购买;同时,天兴仪表向北京贝瑞和康生物技术股份有限公司全体股东发行股份购买其持有的贝瑞和康100%股权。重组完成后,上市公司将持有贝瑞和康100%股权,贝瑞和康借壳天兴仪表,实现A股上市。但关于具体的重组方案并未披露。

    2016年12月

    天兴仪表披露重大资产重组草案,拟以21.14元/股,发行股份2.03亿股,作价43亿元购买贝瑞和康100%股权。同时向大股东天兴集团的控股子公司,2.97亿元出售上市公司资产与负债。交易完成后,基因测序企业贝瑞和康将借壳上市。同时披露2013年、2014年、2015年和2016年上半年,贝瑞和康主营业务收入分别为2.58亿元、3.34亿元、4.45亿元和3.85亿元,增长态势良好。同期,医学产品及服务业务贡献的毛利分别为1.6亿元、2.08亿元、2.38亿元和2.43亿元,占贝瑞和康主营业务毛利额的比例分别为90.51%、89.04%、88.63%和97.29%,为贝瑞和康利润的最主要来源。业绩补偿方承诺2017年、2018年、2019年的净利润分别不低于2.284亿元、3.092亿元和4.045亿元。天兴仪表股票于12月19日复牌后,迎来连续10个涨停板。

    2017年1月

    证监会向天兴仪表寄出《行政许可项目审查一次反馈意见通知书》,并对此次借壳重组事项连续提出33个问题,要求天兴仪表作出书面说明和解释,并在30个工作日内向证监会提交书面回复意见。

    2017年2月

    天兴仪表股价创出了近一个月的新高46.86元。2月28日,该股票被停牌一天。同时,该公司因 2015年、2016年连续两个会计年度经审计的净利润为负值,根据《深圳证券交易所股票上市规则》的相关规定,深圳证券交易所有权对公司股票交易实行“退市风险警示”。股票简称由“天兴仪表”变更为“*ST天仪”,实行退市风险警示后公司股票交易的日涨跌幅限制为 5%。

    2017年3月

    *ST天仪(000710)于3月9日发布了一份长达131页篇幅,约十万字的答复书,公布了一系列有关贝瑞和康的相关数据,包括近些年销售业绩、与主要竞争对手的基本情况对比分析,以及答复证监会此前下发的《中国证监会行政许可项目审查一次反馈意见通知书》等内容。3月21日,证监会又向天兴仪表寄出《行政许可项目审查二次反馈意见通知书》,并对此次借壳重组事项继续提出3个问题。

    2017年4月

    4月12日,*ST天仪控股股东天兴集团所持有的44,002,000股股份(占总股本的29.10%)被四川省高级人民法院冻结,并于4月22日解除冻结。期间,为了回复证监会的二次反馈意见,天兴仪表再次调整贝瑞和康重组方案,将鼎锋明德致知、鼎锋明德正心已经将其所持贝瑞和康1,563,672股股份全部转让给君联茂林、苏州启明创智、鼎锋海川。4月20日,证监会发布《并购重组委2017年第19次工作会议公告》,定于2017年4月26日上午9:00召开2017年第19次并购重组委工作会议,其中审核的并购重组申请人包括成都天兴仪表股份有限公司。4月26日,证监会上市公司并购重组审核委员会召开了第19次会议,审核通过了成都天兴仪表股份有限公司发行股份购买资产的相关事宜,标志着基因测序领先企业贝瑞和康借壳上市取得了实质性的成功!

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    Amresco 0706蛋白酶 K

    Amresco 0706蛋白酶 K
    中文名称:蛋白酶K

    英文名称:Proteinase K from Tritirachium album
    CAS号:39450-01-6
    级别:Chromatographically purified
    分子量:~18kDa
    Activity (hemoglobin,pH 7.5; 37 °C):≥ 30mAnsonU/mg
    Specific activity:≥40mAnsom u/mg protein
    DNases (Nicking activity,pBR 322,6 h,37℃):Not detectable
    RNases (RNA,2 h,37℃):Not detectable
    Solubility in water:(20 ℃) soluble
    pH value:6.2~6.8 (10 g/l,H2O,20℃)
    蛋白酶K 性状:白色或类白色粉末,是一种非特异性、枯草杆菌蛋白酶相关的丝氨酸蛋白酶, 来源于白色念球菌(. Tritirachium album. ),具有很高的比活性。EDTA等鳌合剂或SDS等去垢剂不能使蛋白酶K失活,反而增强和稳定酶活性,并在较广的PH范围内(PH4-12.5)均有活性。在组织或细胞核酸分离时,使核酸酶(RNA和DNA)以及反应中其他酶失活。
    蛋白酶K 用途:生化研究。广谱蛋白酶,95C 加热10 分钟,则完全失活。可替代 DEPC 处理 RNA 抽提用的离心管和枪头,实现灭活 RNase A 的目的,也用于处理 RNase-Free的水。充分降解组织或细胞中蛋白质分子特别是核酸,以达到释放和萃取高质量核酸的辅助试剂。用于质粒或基因组DNA、RNA的分离以及蛋白多肽印迹实验。
    保存:2~8℃
    蛋白酶K 化学试剂的种类很多,世界各国对化学试剂的分类和分级的标准不尽一致。
    IUPAC对化学标准物质的分类为:
    A级:原子量标准。
    B级:和A 级最接近的基准物质。
    C级:含量为100+-0.02%的标准试剂
    D级:含量为100+-0.05%的标准试剂
    E级:以C级或D级为标准对比测定得到的纯度的试剂
    化学试剂按用途可分为标准试剂,一般试剂,生化试剂等等。
    我国习惯将相当于IUPAC 的C级、D级的试剂称为标准试剂。

    学校供给饮食相关_脂肪残留物性试验用试剂|简易分析|一般分析纯试剂|试剂|关东化学株式会社

    脂肪性残留物试验用红辣椒酒精溶液

    “学校供给饮食法》规定的“学校供给饮食卫生管理标准》中,餐具清洗状态为了调查的定期检查的义务。红辣椒酒精溶液,餐具类残留附着的脂肪成分染色的试剂,肉眼简单地能判别。操作方法也很简单,日常的检查也能使用。

    红辣椒酒精溶液(2016年4月27日更新)

    学校供给饮食相关_脂肪残留物性试验用试剂|简易分析|一般分析纯试剂|试剂|关东化学株式会社

    检验方法

    试剂的咨询

    信息查询

    电话: 03 – 6214 – 1090

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    甘油检测试剂盒 K-GCROL 70 assays (manual) / 700 assays (microplate)

    甘油检测试剂盒
    英文名:Glycerol Assay Kit
    货号:K-GCROL
    规格:70 assays (manual) / 700 assays (microplate)
    市场价: 1808元

    分析物意义:常见食品组分,或作为甜味剂,或用于改善口感

    Megazyme检测试剂盒优点:新型的药片模式,性质更稳定,反应快

    The Glycerol test kit is a simple, reliable, rapid and accurate method for the measurement and analysis of Glycerol in beverages, foodstuffs and other materials.
    Suitable for manual and microplate formats.

    UV-method for the determination of Glycerol in foodstuffs,
    beverages and other materials

    Principle:
    (glycerokinase)
    (1) Glycerol + ATP → L-glycerol-3-phosphate + ADP

    (pyruvate kinase)
    (2) ADP + PEP → ATP + pyruvate

    (L-lactate dehydrogenase)
    (3) Pyruvate + NADH + H+ → L-lactic acid + NAD+

    Kit size: 70 assays (manual) / 700 (microplate)
    Method: Spectrophotometric at 340 nm
    Reaction time: ~ 5 min
    Detection limit: 0.34 mg/L
    Application examples:
    Wine (and grape juice), beer, spirits, vinegar, marzipan, fruit juices,
    soft drinks, toothpaste, honey, tobacco, paper (and cardboard),
    cosmetics, pharmaceuticals, soap and other materials (e.g. biological
    cultures, samples, etc.)
    Method recognition:
    Methods based on this principle have been accepted by OIV and
    MEBAK

    Advantages

    • Novel tablet format for increased stability
    • Very competitive price (cost per test)
    • All reagents stable for > 2 years as supplied
    • Very rapid reaction
    • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
    • Standard included
    • Suitable for manual and microplate formats

     

     Q1. There is an issue with the performance of the kit; the results are not as expected.

    If you suspect that the Megazyme test kit is not performing as expected such that expected results are not obtained please do the following:

    1. Ensure that you have tested the standard sample that is supplied with the Megazyme test kit.
    2. Send the results of the kit standard, blank samples and the results obtained for your sample, in the relevant MegaCalc spreadsheet (if available) to Megazyme (cs@megazyme.com). Where available the relevant MegaCalc spreadsheet can be downloaded from where the product appears on the Megazyme website.
    3. State the kit lot number being used (this is found on the outside of the kit box).
    4. State which assay format was used (refer to the relevant page in the kit booklet if necessary).
    5. State exact details of any modifications to the standard procedure that is provided by Megazyme.
    6. State the sample type and describe the sample preparation steps if applicable.

    Q2. Is K-GCROL specific for glycerol?

    K-GCROL is highly specific for glycerol.
    Some compounds that are known not to react or interfere with the assay include:
    Polyethylene glycol
    Ethylene glycol
    Propylene glycol

    Q3. Sometimes a negative absorbance change is obtained for the blank samples, is this normal? Should the real value (negative absorbance change) or “0” be used in the calculation of results?

    Sometimes the addition of the last assay component can cause a small negative absorbance change in the blank samples due to a dilution effect and in such cases it is recommended that the real absorbance values be used in the calculation of results.

    Q4. Should the pH of the sample be adjusted even for samples in acidic media?

    The pH of the assay solution after the sample is added should be the same as that of the assay buffer that is supplied with the kit.
    Low sample volumes (e.g. 0.1 mL) are not likely to affect the pH of the assay solution and therefore may not require pH adjustment.
    Samples above 0.1 mL are more likely to affect the pH of the assay solution and therefore the pH of these samples should be adjusted as described in the data booklet, prior to addition to the assay.

    Q5. Is the Glycerol Assay Kit (K-GCROL) suitable for measurement using cell culture media samples?

    Yes, assuming that the concentration of the analyte in the sample (after sample preparation) is above the limit of detection for the kit.  It may be sufficient to use the sample directly in the assay after clarification by centrifugation / filtering followed, by dilution (if required) in distilled water.

    Q6. Can the test kit be used to measure biological fluids and what sample preparation method should be used?

    The kit assay may work for biological fluids assuming that inositol is present above the limit of detection for the kit after any sample preparation (if required). Centrifugation of the samples and use of the supernatant directly in the kit assay (with appropriate dilution in distilled water) may be sufficient. However, if required a more stringent sample preparation method may be required and examples are provided at the following link:http://www.megazyme.com/docs/analytical-applications-downloads/biological_samples_111109.pdf?sfvrsn=2

    The test kit has not been tested using biological fluids as samples because it is not marketed or registered as a medical device. This will therefore require your own validation.

    Q7. Can the manual assay format be scaled down to a 96-well microplate format?

    The majority of the Megazyme test kits are developed to work in cuvettes using the manual assay format, however the assay can be converted for use in a 96-well microplate format. To do this the assay volumes for the manual cuvette format are reduced by 10-fold. The calculation of results for the manual assay format uses a 1 cm path-length, however the path-length in the microplate is not 1 cm and therefore the MegaCalc spreadsheet or the calculation provided in the kit booklet for the manual format cannot be used for the micropalate format unless the microplate reader being used can.

    There a 3 main methods for calculation of results using the microplate format:

    1. The easiest method is to use a microplate reader that has a path-length conversion capability (i.e. the microplater reader can detect the path-length of each well and convert the individual readings to a 1 cm path-length). This will allow values to be calculated using the MegaCalc calculation software which can be found where the product is located on the Megazyme website.
    2. Perform a standard curve of the analyte on each microplate that contains test samples and calculate the result of the test samples from the calibration curve (concentration of analyte versus absorbance).
    3. Perform a standard curve of the analyte in both the cuvette format (i.e. with a 1 cm path-length) and the 96-well microplate format and use these results to obtain a mean conversion factor between the cuvette values and the microplate values. Subsequent assays in the microplate format can then be converted from the calculated conversion factor.

    Q8. How can I work out how much sample to extract and what dilution of my sample should be used in the kit assay?

    Where the amount of analyte in a liquid sample is unknown, it is recommended that a range of sample dilutions are prepared with the aim of obtaining an absorbance change in the assay that is within the linear range.
    Where solid samples are analysed, the weight of sample per volume of water used for sample extraction/preparation can be altered to suit, as can the dilution of the extracted sample prior to the addition of the assay, as per liquid samples.

    Q9. The pH of my sample is low (pH ~ 3.0), do I need to adjust this before I use the sample in the kit assay?

    The final pH of the kit assay after the sample is added should not change from what it should be (as stated in the kit for the assay buffer). If it does change then the sample will require pH adjustment. In most cases the sample volume being used is low relative to the final assay volume and in this case the pH of the kit assay is unlikely to be affected.

    Q10. Can you explain, step by step, how to follow the method and perform the kit assay?

    For users who are not familiar with how to use the Megazyme tests kits then it is recommended that they follow this example, e.g. D-Fructose/D-Glucose Assay kit K-FRUGL (http://secure.megazyme.com/D-Fructose-D-Glucose-Assay-Kit):

    1. The kit components are listed on pages 2-3 of the kit booklet.
    2. Prepare the kit reagents as described on page 3.
    3. For separate measurements of glucose and fructose follow procedure A on page 4.
    4. Pipette the volumes listed for water, sample, solution 1 and solution 2 into 3 mL, 1 cm pathlength cuvettes. Duplicate sample assays and duplicate blanks are recommended. Mix the contents of each cuvette by inversion (seal the cuvette using parafilm or a plastic cuvette cap – do not use a finger) then after ~3 min record the first absorbance reading of each cuvette at 340 nm (this is reading A1).
    5. Then add suspension 3 and mix the contents of each cuvette by inversion. Incubate for 5 minutes then record the absorbance reading of each cuvette at 340 nm (this is reading A2). NB. It is essential that the reaction is compete. To assess this, record the absorbances at ~ 2 minute intervals and until the absorbance plateaus. A stable absorbance indicates that the reaction is complete. If the absorbance continues to increase then continue to record absorbances until it plateaus and only then record absorbance reading A2.
    6. Then add suspension 4 and mix the contents of each cuvette by inversion. Incubate for 5 minutes then take absorbance reading of each cuvette at 340 nm (this is reading A3). NB. As above, assess that the reaction has completed by take subsequent readings at ~2 min intervals.
    7. For simple, automated results analysis, input the absorbance readings (A1, A2, A3) for samples and blanks into the K-FRUGL MegaCalc.

    To ensure that the assay is working, and being performed correctly it is recommend that the test is performed using the standard sample that is provided with the kit and to obtain the expected values before proceeding to test real samples.
    It is recommend that new users also watch this video which highlights how to perform the assays.
    Many of the other Megazyme test kits follow a similar format.

    Q11. I have some doubts about the appearance/quality of a kit component what should be done?

    If there are any concerns with any kit components, the first thing to do is to test the standard sample (control sample) that is supplied with the kit and ensure that the expected value (within the accepted variation) is obtained before testing any precious samples. This must be done using the procedure provided in the kit booklet without any modifications to the procedure. If there are still doubts about the results using the standard sample in the kit then send example results in the MegaCalc spread sheet to your product supplier (Megazyme or your local Megazyme distributor).

    Q12. How much sample should be used for the clarification/extraction of my sample?

    The volume/weight of sample and total volume of the extract can be modified to suit the sample. This will ultimately be dictated by the amount of analyte of interest in the sample and may require empirical determination. For low levels of analyte the sample:extract volume ratio can be increased (i.e. increase the sample and/or decrease the total extraction volume).

    Alternatively, for samples with low concentrations of analyte, a larger sample volume can be added to the kit assay. When altering the sample volume adjust the distilled water volume added to the assay accordingly so that the total assay volume is not altered.

    Q13. Can the sensitivity of the kit assay be increased?

    For samples with low concentrations of analyte the sample volume used in the kit assay can be increased to increase sensitivity. When doing this the water volume is adjusted to retain the same final assay volume. This is critical for the manual assay format because the assay volume and sample volume are used in the calculation of results.

    Q14. When using this kit for quantitative analysis what level of accuracy and repeatability can be expected?

    The test kit is extremely accurate – at Megazyme the quality control criteria for accuracy and repeatability is to be within 2% of the expected value using pure analytes.

    However, the level of accuracy is obviously analyst and sample dependent.

    Q15. Is it possible to add a larger volume then 2 μL of enzyme to the microplate assay? In some instances 2 μL can be difficult to pipette manually.

    Yes, instead of adding 2 μL of enzyme suspension an alternative is to dilute the enzyme and add a larger volume to the microplate assay.

    Dilute the assay buffer 10-fold with distilled water and use this as the diluent to dilute an aliquot of the enzyme suspension also by 10-fold. Instead of 2 μL, use 20 μL of the diluted enzyme in the microplate assay.

    Q16. To measure fermentation samples contain high microbial cell density, is cell disruption required?

    Cell disruption is only require to measure glycerol within microbial cells.

    To measure glycerol in the extracellular media only then cell disruption is not required and centrifugation of the sample may be sufficient, e.g.:

    (a) Determination of glycerol in cell culture media/supernatants. In general, the concentration of glycerol in cell culture media/supernatants can be determined without any sample treatment (except clarification by centrifugation/filtering or dilution according to the dilution table, if necessary). Typically, no clarification or dilution is required, and a sample volume of 0.1 mL is satisfactory.

    If interference is suspected then sample clarification/deproteinisation using carrez reagents or perchloric acid should be used (methods are provided in the kit booklet).