T****r
发帖数: 9 | 1 hybridoma一般很容易复苏，但是有时候如果冻存条件不好有可能遇到复苏困难的情况
。细胞往往开始看起来很好，但是根本长不起来。这种情况一般是因为真正有活性的细
胞比例太小的原因。因为hybridoma具有cluster生长的优势，很多paracrine
sufficient的信号有利于生长。如果长不好的话建议用hybridoma cloning factor 加
在culture
media 中，或者加入feeder cells.
10000rpm
了。 |
|
s*******b
发帖数: 1012 | 2 在自己做单抗中，想请教一下版上的大牛，拿到离心过后的hybridoma的supernatant（
没有purification），要如何测量里面IgG的含量？已经有了rabbit IgG whole
molecule可以用作standard。Bradford？BCA？ or some other methods？ Hybridoma
supernatant里的主要蛋白是不是就是分泌的IgG？多谢多谢 |
|
c*****i
发帖数: 1392 | 3 Hybridoma是最容易复苏的一类细胞了。 37C水浴速融，贴壁的Hybridoma直接加入培养
瓶，2小时后换液，可用20％FBS。悬浮细胞解冻后加1ml培养基，慢速离心1000rpm，
5min，去上清，加新鲜medium，培养。若细胞太少，培养一周后看
10000rpm
了。 |
|
S*****a
发帖数: 63 | 4 Boston地区大药厂
现有２个Antibody Engineering/Yeast Display/Hybridoma 工作openings
一个是PhD level;另一个是BS/MS level
欢迎有Antibody Engineering/Yeast Display或Hybridoma背景的朋友申请(须有绿卡或
EAD)。
有意者请Email: j**************[email protected] |
|
p****c
发帖数: 233 | 5 I want to culture a hybridoma line to get antibody. Please give some
suggestions regarding cell culture medium. Should I use less FBS to
facilitate the subsequent purification.
Thanks |
|
f*******g
发帖数: 35 | 6 请教高手一个问题， 我从hybridoma cells 纯化抗体， 得到的抗体 跑胶， 除了重链
（55KD）和轻链 （25KD）外，还有一条 带，在轻链下面。 请问这可能是什么原因造
成的呢？
谢谢了！！！ |
|
f*******g
发帖数: 35 | 7 培养hybridoma cells, 收集上清, 用gamma-bind plus beads (protein G) 纯化 抗体
,然后concentrate elute. buffer change to PBS
谢谢! |
|
c*****i
发帖数: 1392 | 8 你是说Gibco的hybridoma SFM吧，比FBS medium还容易，Protein A or G即可 |
|
l*********2
发帖数: 247 | 9 第一次接触Hybridoma cell。细胞从液氮取出后在70%酒精里融化。培养液加热至37度
，替换冻存液，然后细胞在6孔板中培养。24小时后吸取培养液至2ml tube，10000rpm
离心后复加至细胞。24小时有很多贴壁细胞，48小时观察只有少量贴壁，大部分死掉了。
已重配过培养液，不是培养液的原因。
请问板上高手这是什么原因？谢谢！ |
|
f*****f
发帖数: 195 | 10 Hybridoma不是典型的贴壁细胞，大部分悬浮，不是死细胞。那些贴着的轻微吹打就下
来了。
70%酒精里融化? 直接37度水浴融化。
10000rpm 太高，1000rpm 5 min足够
10000rpm
了。 |
|
|
发帖数: 1 | 12
Download pdf published by SfN (Chapter from 'The History of Neuroscience in
Autobiography, Volume 8' edited by Larry R. Squire)
Yuh-Nung Jan's CV
Lily Jan's CV
Yuh-Nung Jan and Lily Jan
Birth
Family History and Growing Up
National Taiwan University
The Hiking Trip to Shitou in the Spring of 1967
Graduate School Application
Graduate Study at Caltech (1968锟974)
Seymour Benzer Lab (1974锟977)
Steve Kuffler锟絪 Lab at H... 阅读全帖 |
|
y****o
发帖数: 26 | 13 【 以下文字转载自 Biology 讨论区 】
发信人: yayayo (yayayo), 信区: Biology
标 题: 求推荐麻省的技术员工作，公司或学校，7月上班，不要签证
发信站: BBS 未名空间站 (Sat Apr 6 14:41:39 2013, 美东)
本人女，<30岁，有美国的master, 在美国有5年实验室工作经验。因6月底要搬家到麻
省，现在正在找技术员的工作。
我有丰富的动物实验经验，including breeding, genotyping, bleeding, IP
injection, tail vein injection, retro-orbital injections, intragastric
administration, dissection of rats & mice, lethal bone marrow transplant.同
时也擅长细胞培养，包括cell line, primary cell culture, hybridoma cell
culture。常用的bench work 基本都做过，比如FACS，Immun... 阅读全帖 |
|
y****o
发帖数: 26 | 14 【 以下文字转载自 Biology 讨论区 】
发信人: yayayo (yayayo), 信区: Biology
标 题: 求推荐麻省的技术员工作，公司或学校，7月上班，不要签证
发信站: BBS 未名空间站 (Sat Apr 6 14:41:39 2013, 美东)
本人女，<30岁，有美国的master, 在美国有5年实验室工作经验。因6月底要搬家到麻
省，现在正在找技术员的工作。
我有丰富的动物实验经验，including breeding, genotyping, bleeding, IP
injection, tail vein injection, retro-orbital injections, intragastric
administration, dissection of rats & mice, lethal bone marrow transplant.同
时也擅长细胞培养，包括cell line, primary cell culture, hybridoma cell
culture。常用的bench work 基本都做过，比如FACS，Immun... 阅读全帖 |
|
c*o
发帖数: 186 | 15 新工作到一个地方，做Hybridoma,时不时有mold出来，也没有大面积的contaminate，
就是一天一，两个well那种。放别人incubator里面的hybridoma没事儿，我自己
incubator里面别的细胞也没事儿。养细胞也不是新手了，以前从来没有遇到过这种情
况。大家帮我分析分析吧。 |
|
s**u
发帖数: 9035 | 16 Molecular Biologist
Qualifications:
Advanced degree in Molecular or Cell Biology, with 5+ yrs experience or Ph.D
. with 0-2 yrs experience, in genetic engineering;
Excellent attention to detail, good communication skills;
Team oriented, but can perform independently.
The ideal candidate has recent hands-on experience in transferring antibody
genes from hybridomas to cell lines.
Principal Responsibilities:
Isolate genetic DNA from hybridomas, create vector(s) and transfect
monoclonal antibody int... 阅读全帖 |
|
y****o
发帖数: 26 | 17 本人女，<30岁，有美国的master, 在美国有5年实验室工作经验。因6月底要搬家到麻
省，现在正在找技术员的工作。
我有丰富的动物实验经验，including breeding, genotyping, bleeding, IP
injection, tail vein injection, retro-orbital injections, intragastric
administration, dissection of rats & mice, lethal bone marrow transplant.同
时也擅长细胞培养，包括cell line, primary cell culture, hybridoma cell
culture。常用的bench work 基本都做过，比如FACS，Immunohistochemistry， ELISA
， Production and purification of monoclonal antibody from hybridoma cells，
Western blotting, Quantitative real-time... 阅读全帖 |
|
c*****i
发帖数: 1392 | 18 一般而言10％FBS上清可以直接用于western.其实Hybridoma培养可以用serum free
medium.比如Gibco的Hybridoma SFM. |
|
D***a
发帖数: 516 | 19 刚好也要养，有一个问题问一下：hybridoma扩增用RPMI 10%FBS，产抗体用hybridoma
SFM，中间要不要让细胞适应几代，还是扩增到足够细胞，直接换SFM？谢谢。 |
|
s**u
发帖数: 9035 | 20 发信人: shmu (shmu), 信区: Biology
标 题: opening Molecular Biologist position
发信站: BBS 未名空间站 (Wed Jan 25 18:32:38 2012, 美东)
Molecular Biologist
Qualifications:
Advanced degree in Molecular or Cell Biology, with 5+ yrs experience or Ph.D
. with 0-2 yrs experience, in genetic engineering;
Excellent attention to detail, good communication skills;
Team oriented, but can perform independently.
The ideal candidate has recent hands-on experience in transferring antibody
genes from hybridomas to cell lines.
... 阅读全帖 |
|
s*i
发帖数: 5025 | 21 癌细胞，跟正常细胞并没有太大区别，除了一点：就是癌细胞该死的时候不死。老的不
死，新的源源不断，最终导大瘤子。
利用癌细胞，能得奴贝尔奖的。参考 hybridoma |
|
发帖数: 1 | 22 B淋巴细胞就是从脾脏中取出来的
但是没有国家敢越这个雷池，
其实大家都知道答案，包括美国科学家
杂交瘤细胞（hybridoma）是一种在制备单克隆抗体过程中，用骨髓瘤细胞和B淋巴细胞
融合而成的细胞。杂交瘤细胞一般通过瘤细胞培养来制备。 |
|
i********g
发帖数: 77 | 23 Another one at same location:
Job Description
This position is in a group that manufactures intermediates to support
reagent manufacturing in in-vitro diagnostics. The applicant should have a
BS in Chemistry, Biochemistry or related discipline with some experience or
an advanced degree in a similar field. Applicants with some industrial
background and knowledge of GMP/ISO will be preferred.
The successful candidate will have some experience in mammalian cell culture
and hybridoma work.
Knowledge... 阅读全帖 |
|
m****s
发帖数: 18160 | 24 【 以下文字转载自 Returnee 讨论区 】
发信人: RogerMiao (小猎人Roger), 信区: Returnee
标 题: 【招聘海归博士】
关键字: 招聘，海归，抗体，免疫，生物
发信站: BBS 未名空间站 (Thu Nov 28 23:03:14 2013, 美东)
国内顶级生物可以公司招聘海归博士！！
欢迎自荐或者推荐！！
联系邮箱r********[email protected]
Job Title: Senior Scientist position in Antibody department
Job Description：
1、Provide support for premium hybridoma relevant antibody projects,
customer communication from quotation to whole project management.
2、Be involved in internal antibody project design and technical support.
3、Provide super... 阅读全帖 |
|
R*******o
发帖数: 19 | 25 Job Title: Senior Scientist position in Antibody department
Job Description：
1、Provide support for premium hybridoma relevant antibody projects,
customer communication from quotation to
whole project management.
2、Be involved in internal antibody project design and technical support.
3、Provide supervisory and training to associate and assistant scientists.
4、Be responsible for research and development for new technology
development to promote our antibody services and products.
5、Working with m... 阅读全帖 |
|
R*******o
发帖数: 19 | 26 【招聘】【抗体工程】【海归博士】
Job Title: Senior Scientist position in Antibody department
Job Description
1•Provide support for premium hybridoma relevant antibody projects,
customer communication from quotation to whole project management.
2•Be involved in internal antibody project design and technical
support.
3•Provide supervisory and training to associate and assistant
scientists.
4•Be responsible for research and development for new technology
development to promote our antibody... 阅读全帖 |
|
f******1
发帖数: 18 | 27 Nanjing biological company is looking for excellent talents to join it.
（1）抗体高级研究员
Job Description
1·Provide support for premium hybridoma relevant antibody projects,
customer communication from quotation to whole project management.
2·Be involved in internal antibody project design and technical support.
3·Provide supervisory and training to associate and assistant scientists.
4·Be responsible for research and development for new technology
development to promote our antibody services and produ... 阅读全帖 |
|
发帖数: 1 | 28 As a thriving pharmaceutical company, Protheragen Inc. ("Protheragen")
focuses on acquiring, developing and commercializing novel pharmaceutical
and biotechnology products for both therapeutic purposes and diagnostic
applications. Guided by our advanced business strategy – the “VIC R&D
model”: V (risk investment), I (IP right) and C (CRO), we are dedicated to
making drug discovery and development activities the most effective via a
collection of efforts and advantages from all agencies.
We are p... 阅读全帖 |
|
R*******o
发帖数: 19 | 29 猎头岗位，招聘海归博士后
Job Title: Senior Scientist position in Antibody department
Job Description
Provide support for premium hybridoma relevant antibody projects
, customer communication from quotation to whole project management.
Be involved in internal antibody project design and technical
support.
Provide supervisory and training to associate and assistant
scientists.
Be responsible for research and development for new technology
development to promote... 阅读全帖 |
|
c*******y
发帖数: 1657 | 30 Developmental Studies Hybridoma Bank
at the University of Iowa
他们卖的抗体比公司的便宜
可不知道是否有质量保证呢？
如果只买最便宜的上清液
蛋白浓度如何确定呢？ |
|
p*****m
发帖数: 7030 | 31 呃 不过得说明 我们买的都是fly antibody，别的物种的不清楚啊
Developmental Studies Hybridoma Bank
at the University of Iowa
他们卖的抗体比公司的便宜
可不知道是否有质量保证呢？
如果只买最便宜的上清液
蛋白浓度如何确定呢？ |
|
s******r
发帖数: 2876 | 32 跑个胶，染一下，跟标准marker比一下。
Developmental Studies Hybridoma Bank
at the University of Iowa
他们卖的抗体比公司的便宜
可不知道是否有质量保证呢？
如果只买最便宜的上清液
蛋白浓度如何确定呢？ |
|
|
a****d
发帖数: 1919 | 34 We use Mouse IgM anti-O4 from hybridoma bank, and Mouse IgG2b Anti Tuj1 from
Sigma. |
|
j********8
发帖数: 21 | 35 marker跟俺用的一样 其余的不知道：）
帮顶 |
|
V********n
发帖数: 305 | 36 how u purified the ab, more details please |
|
|
V********n
发帖数: 305 | 38 俺用的方法。也是用Protein G的办法，Buffer 用Pierce的Binding Buffer和Elution
Buffer。得到的Elution，直接用Millipore的Spin Column清洗加浓缩，选大一些的Cut
（比如５０KD），避免小片段污染。
看你的胶，有小片段（～２０ KD），可能是蛋白降解，也可能是小片段污染。另，高
分子处还有条带（～２００KD），可检查蛋白变性是否彻底。 |
|
f*******g
发帖数: 35 | 39 没有control, 抗体纯化, 用什么做negative control呢? 我ｂｉｎｄｉｎｇ ｂｕｆ
ｆｅｒ 用的ＰＢＳ， Ｅｌｕｔｉｏｎ ｂｕｆｆｅｒ 用的是 Ａｃｅｔｉｃ Ａ
ｃｉｄ，用ＮａＯＨ中和， 然后用 Ｍｉｌｌｉｐｏｒｅ的Ｓｐｉｎ Ｃｏｌｕｍｎ
清洗 浓缩， ｃｕｔ好像用的是３０ ＫＤ的，
浓缩的时候，很困难！ 需要半小时到一小时的样子，才能浓缩，而且有很多ｐｒｅｃｉｐｉｔａｔｅｓ， 不知道原因？
谢谢楼上的回复，小片段污染常见么？ |
|
|
V********n
发帖数: 305 | 41 呵呵，RE这个。没注意LZ的培养条件，俺一般是从ascites开始提的。 |
|
i********g
发帖数: 77 | 42 Another one at same location:
Job Description
This position is in a group that manufactures intermediates to support
reagent manufacturing in in-vitro diagnostics. The applicant should have a
BS in Chemistry, Biochemistry or related discipline with some experience or
an advanced degree in a similar field. Applicants with some industrial
background and knowledge of GMP/ISO will be preferred.
The successful candidate will have some experience in mammalian cell culture
and hybridoma work.
Knowledge... 阅读全帖 |
|
|
m*p
发帖数: 226 | 44 Only 1-2% of the total IgG in the supernatant are the ones that can react
with your antigen (I can not remember where I read it). |
|
l*****x
发帖数: 658 | 45 Can anyone share the sequences of J and C primers to confirm the presence of
TCR gene in hybridoma fusions?
Thanks |
|
|
M*****n
发帖数: 16729 | 47 这个工作transferring antibody genes from hybridomas to cell lines有什么trick
吗？
.D
antibody |
|
l******g
发帖数: 1623 | 48 如果只是把hybridoma抗体的相应gene搞出来到CHO神马的细胞里面表达，做点研究用的
材料，没啥trick。如果要大量作治疗用的抗体，学问就大了。人猿化，序列优化。。。
trick |
|
|
h******u
发帖数: 602 | 50 是从HYBRIDOMA来的单抗。一直都是收集细胞培养的MEDIUM，直接用做免疫组化和
WESTERN。用几次后就扔掉。
现在想节约点，除了分装小体积，避免多次冻溶，是不是还可以加点proteinase
inhibitor？ 别的招呢？ |
|
