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View additional product information for Imject™ Maleimide PEGylated mcKLH - FAQs (77663)
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我们建议疏水残基占序列中所有残基的50％或更少。确保每五个氨基酸至少有一个带电残基：这通常被认为可以增强肽段的溶解度。肽段（与折叠和掩盖疏水性氨基酸的多肽相比）太小而不能折叠，因此只有少数疏水性氨基酸可使它们不溶解。
其他指南包括：
•含有多个半胱氨酸、甲硫氨酸和色氨酸的肽段可能难以合成。
•一些序列在固相肽段合成或切割中存在问题，最好避免；这些序列包括天冬氨酸-脯氨酸（Asp-Pro）序列或在合成期间在其侧链上需要庞大保护基团的氨基酸片段。
•甘氨酸通常具有良好的抗原性，因为它只有一个氢侧链；这使其可以完全旋转。
请注意，我们提供定制抗体制备服务（https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production.html），包括使用我们专有的抗原分析仪和抗原制备工具（https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production/antigen-profiler-antigen-preparation.html）。
大多数多肽抗原的长度范围为12至16个残基，并且相对容易合成。9个残基或更短的肽段已经成为抗体制备的有效抗原，但是长于16个氨基酸的多肽可能含有多个表位并形成二级结构。超过18个残基的多肽开始出现更多的合成挑战。在合成您的多肽之前，我们建议您使用多肽序列进行BLASTP搜索。这是为了确保多肽与宿主动物中完全不相关的蛋白质中的序列不同源或相同。
请注意，我们提供定制抗体制备服务（https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production.html），包括使用我们专有的抗原分析仪和抗原制备工具（https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production/antigen-profiler-antigen-preparation.html）。
抗体对试剂盒包含捕获抗体、检测抗体、重组标准品和HRP偶联物。每个套组都含有足够的试剂来处理四十个96孔板。可查看按靶标提供的抗体对试剂盒列表（https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-assays-analysis/elisa/antibody-pair-kits.html）。
试剂套组包含捕获抗体、检测抗体、重组标准品、HRP偶联物、TMB底物和终止液。每个套组都含有足够的试剂来处理五个96孔板。试剂套组包含在ELISA试剂盒（https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-assays-analysis/elisa/antibody-pair-kits.html）的主列表中（通过“试剂套组”搜索）。
我们提供超过1000种不同的ELISA试剂盒，靶向> 800种不同的分子。您可以查看按靶标查找 ELISA 试剂盒页面（https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-assays-analysis/elisa/elisa-kits/target-specific-elisa-kits.html）。
HRP受叠氮化钠抑制，因此我们的HRP偶联的抗体不含叠氮化钠。然而，我们的未偶联抗体是以含有叠氮化钠的溶液形式提供。
牛血清白蛋白（BSA）是一种蛋白质稳定剂，叠氮化钠是一种防止细菌生长的防腐剂。添加载体蛋白和防腐剂可增加抗体的功能和寿命。含有BSA和叠氮化物的抗体可以在2-4°C下储存。
通常，抗体可以在室温下放置长达一周而不丧失活性。因此，我们的多种抗体都是在常温下运送的。但是，不推荐在室温或更高温度下长时间储存，在这种情况下我们无法保证抗体的性能。如果您还有关于抗体稳定性的其他问题，请联系技术支持LifeScience-CNTS@thermofisher.com。
建议使用非无霜冰箱储存抗体，是因为其不会经过自动除霜循环。为了使冷冻室保持无霜状态，冷冻室实际上会定期用加热线圈对其进行除霜。在除霜循环期间，冷冻室内的温度可升高至0°C及以上。如果抗体长时间储存于这些温度下，则会部分解冻。解冻大多发生在样品容器中的空气与冷冻物的表面之间的界面处。由于空气比水升温快，因此界面处的蛋白质比试管深处的蛋白质受到的影响更大。
即使您使用的是非无霜冰箱，也应注意抗体在其中的储存位置。最好将它们存放在冷冻室温度波动最小的区域内，该区域通常位于冰箱后部的中心。不要将抗体储存在冷冻室前部附近或冰箱门的搁架上，因为每次打开冰箱门时，抗体都将暴露于室温下。同时需要确保抗体容器紧密封闭。这有助于防止抗体因升华（冰直接转化为水蒸气，从容器中逸出的）而失水。这种缓慢的过程基本上是在正常大气压下进行冻干，并且可能导致抗体损坏。
请遵循手册和产品标签提供的储存建议。浓缩的抗体可以在2-8°C条件下储存长达1个月。对于长期储存，我们建议将所有抗体等分为一次性使用的量，并在-20°C的非无霜冰箱或-80°C下保存。 避免多次冻融，因为这会降低抗体活性。不要冷冻稀释的抗体。在使用前进行最终的抗体稀释。向抗体中加入诸如甘油的稳定剂（例如，40-50％（v / v））是可行的。但是，除非我们说可以这样做，否则我们无法保证添加稳定剂的产品的性能。
抗体的推荐使用量基于细胞或组织裂解物的蛋白质浓度。此外，用量取决于抗体来源，具体如下：对于每200-500μg细胞或组织裂解蛋白，我们建议添加1-10μg纯化的单克隆或多克隆抗体、1-5μL未纯化的多克隆抗血清、0.2-1μL腹水或20-100μL杂交瘤上清液。这些建议仅供参考，因此您可能需要根据经验优化您的IP反应，以确定产生最高产量和最低背景的抗体量。在大多数情况下，您可以通过进行SDS-PAGE，然后对免疫沉淀物进行蛋白质免疫印迹分析，以及使用抗原特异性抗体染色来实现这一目的。
请记住，如果抗体在免疫组织化学或免疫荧光染色中有效，那么它可能在IP中也起作用。这是因为这些应用都基于抗体对天然蛋白质的抗原识别。判断的唯一方法是使用上面讨论的指南作为起点在IP中尝试使用抗体。
二级抗体可能过于特异（例如，仅识别一抗的一种宿主物种）或过于广谱（例如，识别完整的IgG及其任何片段）。在大多数情况下，通过细心设计实验体系并选择合适的二抗，可以克服这些限制。以下注意事项有助于选择合适的二抗：
1.确定一抗的宿主物种（例如，小鼠抗微管蛋白抗体）。
2.为二体选择合适的宿主物种 — 您需要在与一抗的宿主物种不同的物种中产生的二抗（例如，羊抗小鼠IgG）。
3.考虑二抗的交叉反应性或特异性问题。
◦高度交叉吸附 — 在多标记应用或使用内源性抗体样品时使用
◦特异性 — 与一抗的正确片段、种类或链结合
4.使用适当的检测或纯化方法。
◦标记 — 与正确的酶、标签或荧光基团适当偶联以用于所选择的检测方法
◦与蛋白A、蛋白G或蛋白L结合的能力 — 确保所选的二抗对上游或下游使用的分子具有足够的亲和力（如蛋白A包被的微孔板）
5.考虑所提供的二抗的要求。
◦供应的状态 — 无菌液体或冻干粉，悬浮于PBS或Tris缓冲液中，含有载体蛋白如明胶或白蛋白还是添加了蔗糖或微生物抑制剂等稳定剂
二抗是针对一抗的物种的抗体。因此，您将需要在与一抗的宿主不同的物种中产生的二抗。例如，如果您的一抗为兔源性抗体，则需要在兔以外的宿主物种（例如山羊，小鼠等）中产生的抗兔二抗。
Zymed、Biosource和Caltag曾经销售的许多抗体仍然可以从我们这里购买。大多数情况下，这些产品仍然使用之前的货号。事实上，我们仍在使用这些原始供应商开发的生产方法。这意味着您所习惯的抗体规格、配方、浓度、使用指南和结果仍然与过去相同。已经停产的抗体因以下三个原因被淘汰：1）我们不希望同一种抗体有多个品牌名称；2）我们无法保证产品的可靠供应；或者3）一些抗体的年销量对我们来说太少了。如果您发现文献中引用了Zymed、Biosource或Caltag抗体，但是您不确定是否仍然可以购买，请联系技术支持：LifeScience-CNTS@thermofisher.com。如果技术支持可以明确地识别产品，则很容易确定其是否可以购买。
是的，我们的一些未偶联抗体提供500μL的供货形式。货号为：R950-CUS（抗Myc抗体）和R960-CUS（抗V5抗体）。
缩写PAD代表“多克隆抗体名称（Polyclonal Antibody Designation）”。PAD类似于单克隆抗体的克隆名称或编号，但它被用作多克隆抗体的唯一标识符。PAD是商定的命名系统的一部分，该系统的开发使得研究人员可以在出版物中引用抗体而无需使用货号。
多克隆抗体是从先前免疫的动物的血清中收获的相关抗体的混合物。血清来源的多克隆抗体几乎可以在任何哺乳动物中产生。然而，大多数多克隆抗体是在具有大量血液的动物中产生的，如山羊、兔、驴和绵羊。
多克隆抗体的替代来源是来自已经用抗原免疫的鸟类（如鸡和火鸡）的蛋黄。从蛋黄中分离的多克隆抗体属于IgY类。IgY抗体在功能上等同于来自哺乳动物的IgG抗体。
我们的大多数多克隆抗体产品来自山羊或兔血清。我们提供许多识别众多蛋白质抗原（包括磷酸化靶标）的多克隆抗体。我们的多克隆抗体也包括种类和物种特异性抗体（例如，识别来自特定物种的IgG的抗体），因此它们用作免疫检测的二抗。
如果要选择多克隆二抗，请使用我们的二抗选择工具（https://www.thermofisher.com/order/genome-database/browse/antibody/sub-type/antibody_secondary）。
通常，对于IgG抗体，重链约为55kDa，轻链约为25kDa。
在小鼠、大鼠、仓鼠和兔中可产生单克隆抗体。首先，将抗原注入到宿主动物体内启动机体免疫应答。继而大多数操作都是在体外将来自这些宿主的脾细胞与培养的恶性骨髓瘤细胞进行融合。在融合步骤中存活下来的细胞克隆称为杂交瘤。杂交瘤因骨髓瘤特性而可以永生，且容易在培养物中繁殖。一些杂交瘤克隆因带有B细胞特性而持续合成并分泌针对单表位的遗传学同源性抗体，称为单克隆抗体。单克隆抗体与小鼠、大鼠、仓鼠和兔的天然免疫球蛋白同源，但可由杂交瘤在体外无限生产。小鼠、大鼠、仓鼠和兔源性杂交瘤是目前最常见的单克隆抗体来源，我们提供来自这些物种的单克隆抗体，可识别大量抗原。
请转到我们的一抗选择工具（https://www.thermofisher.com/order/genome-database/browse/antibody/sub-type/antibody_primary），然后在左侧导航栏的“抗体类型”下选择“单克隆”或“重组”，查看我们的小鼠和其他单克隆抗体，或我们的重组ABfinity兔单克隆抗体。
以1:200稀释抗体意味着将1体积的抗体加入到199体积的稀释液中（1 + 199 = 200）。同样地，以1:1000稀释抗体意味着将1体积的抗体加入到999体积的稀释液中（1 + 999 = 1000）。假设您需要20 mL稀释度为1:200的抗体。取1 mL抗体并将其加入199 mL的稀释液中，这可能非常容易。但是，您总共只需要20 mL稀释的抗体，因此您可以取0.1 mL（100μL）抗体并将其加入19.9 mL的稀释液中（0.1 + 19.9 = 20）。所得的溶液仍然是1:200的稀释度。如果您只需要总共2 mL的稀释的抗体，那么混合液将是0.01 mL（10μL）抗体加入1.99 mL的稀释液中（0.01 + 1.99 = 2）。所得的溶液仍然是1:200的稀释度。稀释抗体时，轻轻混合以确保溶液均匀。我们建议不要使用涡旋混合器，因为涡旋可能会导致抗体失活。
我们的大多数抗体产品的浓度都列在产品标签和手册中。对于一抗和二抗，浓度范围为0.1至2 mg/mL，具体浓度因产品而异。大多数“预稀释”抗体的浓度范围为1-10μg/ mL；对于我们称之为“浓缩物”的产品，其浓度通常为50-500μg/ mL。对于一些产品，我们不提供浓度 — 这些产品按体积（例如，100μL）或以其他单位（例如10次印迹）包装。此外，我们也没有提供许多前Zymed病理学抗体（货号以08、18或28开头）的浓度。我们的一些抗体是由未分级血清、浓缩的杂交瘤上清液或腹水样品组成的。在这些情况下，抗体浓度是未知的。对于这些产品，请遵循我们的稀释建议。
按体积包装的抗体浓度不是商业秘密，但其单位大小是基于活性滴度而不是抗体浓度。因此，抗体浓度会有批间差异。对于按体积或其他单位销售的大多数抗体，每批中每个小瓶可以染色的载玻片或蛋白质免疫印迹的数量不会改变，因为推荐的稀释度保持不变。如果您不知道您抗体的浓度，请通过LifeScience-CNTS@thermofisher.com联系技术支持，并提供货号和批号。
We recommend that hydrophobic residues comprise 50% or less of all the residues in your sequence. Make sure there is at least one charged residue for every five amino acids: this is generally known to enhance the solubility of the peptide. Peptides (compared to polypeptides, which fold and bury the hydrophobic amino acids) are too small to fold, so just a few hydrophobic amino acids may leave them insoluble.
Other guidelines include:
Peptides containing multiple Cys, Met, and Trp can be hard to synthesize.
Some sequences are problematic in solid-phase peptide synthesis or cleavage and are best avoided; these include Asp-Pro sequences or stretches of amino acids that require bulky protecting groups on their side chains during synthesis.
Glycine is often good for antigenicity as it has only a hydrogen side chain; this allows for complete rotation.
Please note that we offer a Custom Antibody Production service (https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production.html) that includes the use of our proprietary Antigen Profiler and Antigen Preparation tool (https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production/antigen-profiler-antigen-preparation.html).
Most peptide antigens range in length from 12 to 16 residues and are relatively easy to synthesize. Peptides of 9 residues or shorter have been effective antigens for antibody production, but peptides longer than 16 amino acids may contain several epitopes and form secondary structures. Peptides in excess of 18 residues begin to present more synthetic challenges. Before you synthesize your peptide, we recommend doing a BLASTP search using your peptide sequence. This is to make sure that the peptide is not homologous or identical to a sequence in a completely unrelated protein in the host animal.
Please note that we offer a Custom Antibody Production service (https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production.html) that includes the use of our proprietary Antigen Profiler and Antigen Preparation tool (https://www.thermofisher.com/us/en/home/life-science/antibodies/custom-antibodies/custom-antibody-production/antigen-profiler-antigen-preparation.html).
Improper storage of antibodies can lead to:
- Degradation: Loss of activity and specificity
- Aggregation: Formation of precipitates or clumps
- Reduced performance: Poor results in assays or experiments
Find additional tips, troubleshooting help, and resources within our Antibodies and Immunoassays Support Center.
Antibody pair kits contain capture antibody, detection antibody, recombinant standard and HRP conjugate. Each contains enough reagents to process forty 96-well plates. A list of Antibody Pair Kits (https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-assays-analysis/elisa/antibody-pair-kits.html) is available by target.
Find additional tips, troubleshooting help, and resources within our Antibodies and Immunoassays Support Center.
Reagent Sets contain Capture Antibody, Detection Antibody, Recombinant Standard, HRP Conjugate, TMB Substrate and Stop Solution. Each contains enough reagents to process five 96-well plates. Reagent Sets are included in the main list of ELISA (https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-assays-analysis/elisa/antibody-pair-kits.html) kits (search by “Reagent Set”).
Find additional tips, troubleshooting help, and resources within our Antibodies and Immunoassays Support Center.
Over 1000 different ELISA kits targeting greater than 800 different molecules are available. A search page (Find ELISA Kits by Target - https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-assays-analysis/elisa/elisa-kits/target-specific-elisa-kits.html) is available.
Find additional tips, troubleshooting help, and resources within our Antibodies and Immunoassays Support Center.
Antibodies prepared against a short peptide sequence may not always recognize the full-length protein in which that peptide sequence is located. The peptide sequence only represents a small portion of the entire protein, and the full-length protein is usually a more complex structure with folds, alpha-helices, beta-sheets, and other structural motifs as well as various posttranslational modifications, any of which can shield the epitope from the antibody. This is one reason it is important to read the manuals for our antibodies. They describe the capabilities and specificity of our antibodies and list the applications for which these products are validated.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
In all likelihood, your antibody stopped working because it lost its reactivity. This happens because antibodies (and most other proteins) are less stable at low concentrations (e.g., the µg/mL range and lower) than they are at higher concentrations. For example, proteins adsorb to surfaces like the walls of their containers due to charge-mediated and hydrophobic interactions. This occurs regardless of the protein concentration, and it usually results in some degree of protein denaturation and loss of activity. However, at low protein concentrations the impact of adsorption is larger per unit of time than at higher concentrations. Antibodies in solution also aggregate with each other for the same reasons that they adsorb to surfaces, sometimes resulting in loss of activity. How fast your diluted antibody loses activity in storage is unpredictable, so store your diluted antibodies no longer than overnight at 2-8 degrees C and then discard them. Better yet, make a fresh working dilution each time you need to use the antibody.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
The definition of “didn't work” depends on what you did with the antibody. For example, it could mean that you observed no specific immunofluorescence staining at the antibody concentration that we suggested. It could also mean that, at the antibody dilution we recommended, your western blots showed high background staining. The suggested antibody concentrations specified in our manuals should be considered starting points for further experimentation. These values were derived in our labs during development of the antibody, or they are based on the experiences of our collaborators as well as other customers. If our suggestions don't work for you, try your method again with a different concentration/dilution. Continue optimizing until your results are as good as you can make them. If your experiments still don't “work”, please contact Technical Support at techsupport@thermofisher.com for further help.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
For most of the antibodies that we sell, a single freeze-thaw cycle should not damage them. Remove the antibody from the freezer and thaw it quickly. Mix it gently (do not vortex) and keep it on ice or in a refrigerator at 2-8 degrees C. It can be stored this way for approximately 1 month. If your working dilution of antibody got frozen, it should not be reused after it thaws. Antibodies in diluted solutions are not stable to freezing and thawing.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
HRP is inhibited by sodium azide, so our HRP-conjugated antibodies do not contain this agent. Our unconjugated antibodies are, however, supplied in solutions containing sodium azide.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
Bovine serum albumin (BSA) is a protein-stabilizing agent, and sodium azide is a preservative that prevents bacterial growth. The carrier protein and preservative are added to increase functionality and longevity of the antibodies. Antibodies supplied with BSA and azide can be stored at 2-4 degrees C.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
Generally, antibodies can be left at room temperature for up to a week without loss of activity. Hence, we ship many of our antibodies at ambient temperature. However, longer storage at room temperature or higher is not recommended, and we can't guarantee the performance of the antibody under such circumstances. Please contact Technical Support at techsupport@thermofisher.com if you have further questions about antibody stability.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
Non-frost-free freezers are recommended for antibody storage because they do not go through automatic defrost cycles. To keep a freezer compartment frost-free, the freezer actually defrosts itself periodically with heating coils. Temperatures inside the freezer can rise to 0 degrees C and above during the defrost cycle. If antibodies are subjected to these temperatures for a long enough time, they can undergo a partial thaw. This happens especially at the interface between the air in the sample container and the surface of the frozen contents. Since air warms faster than water, proteins at the interface are affected more than those in the depths of the tubes.
Even if you are using a non-frost-free freezer, you should pay attention to where your antibodies are stored inside it. It is best to store them in an area of the freezer compartment that experiences the least temperature fluctuation, which is usually in the center in the back of the freezer. Do not store antibodies near the front of the freezer compartment or on a shelf in the freezer door, where they are exposed to room temperature every time the freezer door is opened. Make sure as well that the antibody container is tightly sealed. This can help to prevent water loss from the antibody via sublimation (conversion of ice directly to water vapor that escapes from the containers). This slow process is basically freeze-drying at normal atmospheric pressure, and it can lead to damage to the antibody.
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We recommend following the storage recommendations provided in the manual and on the product label. In general, concentrated antibodies in solution can be stored at 2-8 degrees C for up to 1 month.
For long-term storage, we recommend that antibodies be aliquoted into small, single-use amounts and stored at -20 degrees C in a non-frost-free freezer or at -80 degrees C. Avoid multiple freeze-thaw cycles as this will degrade the antibody over time. Make the final working dilution just prior to use and do not freeze diluted antibodies. Additionally, ensure that the storage buffer is compatible with the intended application (e.g., avoid using sodium azide for cell culture application).
Note: Fluorescent conjugated antibodies should not be stored frozen but always stored at 2-8 degrees C.
It is possible to add stabilizing agents like glycerol (40-50% (v/v), ethylene glycol, or sodium azide to the antibody solution. However, unless we say that it is okay to do so, we cannot guarantee the performance of the product with these stabilizing agents added.
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The recommended amounts of antibody to use are based on the protein concentration of the cell or tissue lysate. Also, amounts vary depending on the antibody source, as follows: for each 200-500 µg of cell or tissue lysate protein, we suggest adding 1-10 µg of purified monoclonal or polyclonal antibody, 1-5 µL of an unpurified polyclonal antiserum, 0.2-1 µL of ascites fluid, or 20-100 µL of hybridoma supernatant. These suggestions are guidelines only, so you will probably have to optimize your IP reactions empirically to determine the amount of antibody that provides the best yield and lowest background. Most likely you will do this by performing SDS-PAGE followed by western blotting of the immunoprecipitates and staining with antigen-specific antibodies.
It is useful to remember that an antibody will probably work in IP if it works well in immunohistochemical or immunofluorescence staining. This is because these applications depend on antibody recognition of the antigen in the native protein. The only way to be certain is to try the antibody in IP using the guidelines discussed above as a starting point.
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Secondary antibodies may be either too specific (e.g., recognize only one host species of primary antibody) or too general (e.g., recognize whole IgG and any fragments thereof). In most cases, these limitations can be overcome by carefully designing the experimental system and choosing the appropriate secondary antibody. The following considerations are useful to help choose a secondary antibody:
Determine the host species of the primary antibody (e.g., mouse anti-tubulin).
Select an appropriate host species for the secondary antibody-you will need a secondary antibody that is raised in a species different from the host species of the primary antibody (e.g., goat anti-mouse IgG).
Consider cross-reactivity or specificity issues of the secondary antibody.
Highly cross-absorbed-for multiple-labeling applications or when using samples with endogenous antibodies.
Specificity-binds to correct fragments, classes, or chains of the primary antibody.
Use an appropriate detection or purification method.
Label-appropriately conjugated to the correct enzyme, tag, or fluorophore for the chosen detection method.
Ability to bind to Protein A, Protein G, or Protein L-make sure the secondary antibody chosen has sufficient affinity for the molecules used upstream or downstream (i.e., Protein A-coated microplates).
Consider requirements of the supplied secondary antibody.
Supplied state-sterile liquid or lyophilized, suspended in PBS or Tris buffer, contains carrier proteins such as gelatin or albumin or the addition of stabilizers such as sucrose or microbial inhibitors.
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Secondary antibodies are directed against the species of the primary antibody. Therefore, you will need a secondary antibody that is raised in a species different from the host species of the primary antibody. For example, if your primary antibody is raised in a rabbit, you will need an anti-rabbit secondary antibody raised in a host species other than rabbit (e.g., goat, mouse, etc.).
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Many of the antibodies that were sold by Zymed, Biosource, and Caltag are still available from us. In most cases, the same catalog numbers are still being used. In fact, the same production methods developed by these original suppliers are still used as well. This means that the antibody specifications, formulations, concentrations, usage guidelines, and the results you are accustomed to are the same now as they were in the past. The antibodies that have been discontinued were eliminated for 3 basic reasons: 1) we did not want multiple brand names on the same antibody; 2) we could no longer assure a reliable supply of the products; or 3) some antibodies simply sold too few units per year for us to continue to sell them. When you find a Zymed, Biosource, or Caltag antibody cited in the literature and you're not sure if it's still available, please contact Technical Support at techsupport@thermofisher.com. If Technical Support can positively identify the product, it's easy to determine its availability.
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Yes, a few of our unconjugated antibodies can be purchased in 500 µL volumes. The catalog numbers are: R950-CUS (anti-Myc antibody) and R960-CUS (anti-V5 antibody).
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The abbreviation PAD stands for “Polyclonal Antibody Designation”. The PAD is analogous to the clone name or number for monoclonal antibodies, but it is used as a unique identifier for polyclonal antibodies. The PAD is part of an agreed-upon nomenclature system that was developed so that researchers can cite antibodies in publications without using catalog numbers.
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Polyclonal antibodies are mixtures of related antibodies harvested from the serum of previously immunized animals. Serum-derived polyclonal antibodies can be produced in virtually any mammal. However, most polyclonal antibodies are produced in animals with large blood volumes like goats, rabbits, donkeys, and sheep. An alternative source of polyclonal antibodies is egg yolk from birds like chickens and turkeys that have been immunized with antigens. Polyclonal antibodies isolated from egg yolk are of the IgY class. The IgY antibodies are functionally equivalent to IgG antibodies from mammals. Most of our polyclonal antibody products are derived from goat or rabbit serum. We offer many polyclonal antibodies that recognize numerous protein antigens, including phosphorylated targets. Among our polyclonal antibodies are class- and species-specific antibodies (e.g., those that recognize IgG from a particular species), so they are used as secondary antibodies for immunodetection.
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To select a polyclonal secondary antibody, please use our Secondary Antibody Selection Tool (https://www.thermofisher.com/order/genome-database/browse/antibody/sub-type/antibody_secondary).
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In general, for IgG antibodies, the heavy chain is approximately 55 kDa and the light chain is approximately 25 kDa.
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Monoclonal antibodies are produced in mice, rats, hamsters, and rabbits. First, the host animal is injected with an antigen to initiate a humoral immune response. In most procedures, spleen cells from these hosts are fused in vitro with cultured malignant myeloma cells. The cell clones that survive the fusion step are known as hybridomas. Hybridomas are immortal because of their myeloma characteristics, and they are easily propagated in culture. Because of their B cell properties, some hybridoma clones continually synthesize and secrete a single, genetically homogeneous type of antibody, the monoclonal antibody. Monoclonal antibodies are therefore homologous to natural immunoglobulins from mice, rats, hamsters, or rabbits, but they can be produced by hybridomas in vitro, indefinitely. Mouse-, rat-, hamster-, and rabbit-derived hybridomas are currently the most common sources of monoclonal antibodies, and we offer monoclonals from these species that recognize a huge selection of antigens.
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Please go to our Primary Antibody Selection Tool (https://www.thermofisher.com/order/genome-database/browse/antibody/sub-type/antibody_primary) and then select “Monoclonal” or “Recombinant” under “Antibody Type” on the left navigation pane, for our mouse and other monoclonal antibodies, or our recombinant ABfinity rabbit monoclonal antibodies.
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Diluting an antibody 1:200 means that you add 1 volume of it to 199 volumes of a diluent (1 + 199 = 200). Similarly, a 1:1,000 dilution means that you add 1 volume of antibody to 999 volumes of the diluent (1 + 999 = 1,000). Let's say that you need 20 mL of your antibody at a dilution of 1:200. It would be very easy to take 1 mL of antibody and add it to 199 mL of diluent. However, you only need a total of 20 mL of diluted antibody, so you can take 0.1 mL (100 µL) of antibody and add it to 19.9 mL of diluent (0.1 + 19.9 = 20). This is still a 1:200 dilution. If you only need 2 mL of diluted antibody, the mixture would be 0.01 mL (10 µL) of antibody added to 1.99 mL of diluent (0.01 + 1.99 = 2). This is also a 1:200 dilution. Whenever you dilute an antibody, mix it gently to ensure a homogeneous solution. We recommend against using a vortex mixer, as vortexing may contribute to inactivation of the antibody.
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The concentration of most of our antibody products will be listed on the product label and in the manual. For primary and secondary antibodies, the concentration ranges from 0.1 to 2 mg/mL, depending on the product. The concentration range for most “prediluted” antibodies is 1-10 µg/mL; for what we refer to as “concentrates”, it is usually 50-500 µg/mL. For some products, we do not provide the concentration-these are packed by volume (e.g., 100 µL) or other unit amounts (e.g., 10 blots). Also, the concentrations of many former Zymed pathology antibodies (Cat. No. starting with 08, 18, or 28) are not provided. A few of our antibodies consist of unfractionated serum, concentrated hybridoma supernatant, or samples of ascites fluid. In these cases, the antibody concentrations are unknown. Please follow our dilution recommendations for these products.
The concentrations of antibodies packed by volume are not trade secrets, but the unit size is based on activity titration rather than antibody concentration. Thus, there is lot-to-lot variability in the antibody concentration. For most antibodies sold by volume or other amounts, the number of slides or western blots that you can stain with each vial from each lot does not change because the recommended dilutions remain the same. If you don't know the concentration of your antibody, please contact Technical Support at techsupport@thermofisher.com with the catalog and lot numbers.
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