g***m
发帖数: 465 | 1 Peter Quail's group from Berkley employs light-wavelength sensitive proteins
(phytochrome) to obtain a potential light switched gene expression tool.
In plants, phytochromes exist in 2 states: Pr (sensitive to red light and turn
into Pfr), Pfr (receive far red light and turn into Pr). Only Pfr can bind to
a protein called PIF3.
Now they hook one half of GAL4 to phytochrome, another half to PIF3. Thus only
certain light could cause Pfr binding with PIF3, therefore bring together 2
halves of GAL4, |
|
J**n
发帖数: 1224 | 2 The emmission wavelength of JC-1 is 590, it is orange (for most people).
For ideal flow cytometry, you have to change your filter system to get
optimal result. For confocal microscopy, try to put in approiate filter
(usually it's hard), or you might need to do some real good compensation
(sometime
difficult, too). So, the third choise for confocal will be monitoring only one
channel (decrease of orange or increase of green)
If the nuclear is stained, that means your JC-1 concentration in the cel |
|
s******y
发帖数: 28562 | 3 For detection of protein peak you can use UV absorbance at 260nm. Most
protein has intrinsic absorbance at this wavelength.
For the column, since your protein is big, you can use either Sephedex G-100
(usable for proteins of 40~150kD) or Sephedex G-150 (usable range up to
300kD)or any gel filtration column that can fit your protein.
Since you are not familiar with the chromatophraphy, I suggest you to
collaborate with a protein lab and ask them to do it for you, or at least
assist you to do it. |
|
m**z
发帖数: 787 | 4 invitrogen molecular probes website?
资料 |
|
w********y
发帖数: 288 | 5 一般找到数据也要在自己机器上试一试，有10-20 nm偏差属于正常
资料 |
|
|
i*o
发帖数: 202 | 7 首先感谢大家的回复！
我试了但是只能是单纯地用1photon excitation x 2 作为估计，发现荧光非常弱。
我的样本是这几种beads的混合，所以
不知道看到的都是哪些……
invitrogen 的数据上好像没有这几种，我不熟 不过知道PE－／APC－之后的dye 跟没
有PE－／APC－的在 1photon
excitation上就不一样，不知道2Photon会怎样呢。
所以大家建议我从700 到 1000 nm 试是么？？ |
|
|
s*****j
发帖数: 6435 | 9 2P的光谱和环境关系很大, 各个实验室测的最佳波长相差悬殊.
invitrogen上数据即使有也是放在容剂里测的, 没用.
经验公式是 1photon excitation x 2 - 30nm
700-1000 每隔20nm测一下, 用不了多少时间. |
|
p*******n
发帖数: 12 | 10 Syber red, then use green filter at 523nm wavelength to check the result.
You can buy this dye in invitrogen, about $70 for 50 loadings. |
|
o****d
发帖数: 5454 | 11 that's much different radiation, p32 is β-ray, there is no β-ray in sun's
spectra.
Radiation damage depend on dose and what kind of radiation it is.
β-ray,x-ray and gamma ray are much more dangerous than ultraviolet, etc.
Even low power laser which wavelength is on the visible region like we get
from sun every day also hurt.
some scientist get skin cancer even by their low energy laser in the lab,
and originally when they get the weak laser beam on skin they didn't feel
hurt at all, but after ma... 阅读全帖 |
|
x*******i
发帖数: 32 | 12 Hello there:
Trytophan fluorescence spectroscopy could be used to analyze whether protein
is compact or open. Ex at 288 nm and em around 320 nm suggests that protein
is compact (trytophan in hydrophobic environment). Em at longer wavelength
suggests an open structure. Also relative fluorescence intensity could tell
something? Anybody know why?
Thanks |
|
h*********s
发帖数: 34 | 13 furo2 is ratiometric, which can be used to measure the Ca(II) concentrations.
fluro 3 is intensity based, which can only be used to show the relative
changes of Ca(II).
Also you should check if your fluorescent microscope can be used to read the
two excitation wavelength simultaneously for furo2. |
|
J*****i
发帖数: 55 | 14 Does anyone know whether APC or Alexa Fluor 647 work in Two-photon system?
What is the excitation wavelength? I cannot find any information about this. |
|
s*****j
发帖数: 6435 | 15 just go through the wavelength range of your pulse laser by 10nm step,
then you will know which is best.
two photon excitation spectral curve is normally very broad, the
excitation "peak" doesn't mean much here.
this. |
|
p*******r
发帖数: 4048 | 16
Well, laser has different peak power at different wavelengths. So both need
to be
taken into consideration. |
|
l********l
发帖数: 130 | 17 靠microscope或显微成像得炸药奖的：
The 1925 Nobel Prize in Chemistry and the 1953 and 1986 Nobel Prizes in
Physics are all awarded for the development of different kinds of
microscopes.
1903 – Richard Zsigmondy develops the ultramicroscope and is able to study
objects below the wavelength of light.
The Nobel Prize in Chemistry 1925 was awarded to Richard Zsigmondy "for his
demonstration of the heterogenous nature of colloid solutions and for the
methods he used, which have since become fundamental in modern... 阅读全帖 |
|
y**u
发帖数: 7459 | 18 enzymatic assay 的 color reagent, 说 ‘measure at ex530/em580'.
microplate reader 的 wavelength Lm 该用530还是580？。。。反正我两个都测了一
遍。
谢谢！ |
|
y****6
发帖数: 196 | 19 I am not an expert. But I guess autofluorescence depends on the ex/em
spectra, lower wavelength (blue/green) has higher autofluorescence. |
|
z****u
发帖数: 1007 | 20 同样的一抗拿488，568都没有问题。
就是对excitation wavelength很迷惑。
我们的fluorescent microscope上面3个filter 对应DAPI, FITC 和 TRITC。能否用
DAPI filter用于405，423？ 照理该看到蓝色或者蓝绿色的signal啊。
另外对于647，是不是肉眼无法直接看到？ |
|
z****u
发帖数: 1007 | 21 同样的一抗拿488，568都没有问题。
就是对excitation wavelength很迷惑。
我们的fluorescent microscope上面3个filter 对应DAPI, FITC 和 TRITC。能否用
DAPI filter用于405，423？ 照理该看到蓝色或者蓝绿色的signal啊。
另外对于647，是不是肉眼无法直接看到？ |
|
l**********1
发帖数: 5204 | 22 For LIVE IMAGING
please transfer to LSM 5 Live; Carl Zeiss instrument if your college/university has it.
那本圣经 free download ok now
cited from its pp152
>
A new type of slit-scanning confocal microscope
(LSM 5 Live; Carl Zeiss, Inc.) has
recently been introduced that allows images to
be acquired at rates as fast as or faster than
can be achieved with a spinning-disk confocal
microscope, and with as low or lower rates
of photobleaching. The system adopts principles
from both the spot scanner and ... 阅读全帖 |
|
g*********5
发帖数: 2533 | 23 just stain live cells and wavelength outside of GFP, YFP. Thanks. |
|
f*******e
发帖数: 628 | 24 DsRed 的 spectra 看上去 excitation 的确很 broad, 具体取决于你用的显微镜的
blue 和 green 的波段定义，但 DsRed 在一般蓝光绿光 490nm, 515nm 的地方都有不
是太弱的激发，是最大的 40% 和 70%，信号强的话完全可以引起足够观察到的信号。
然后你们那里的荧光显微镜 emission filter 可能用的是 long pass filter, 就是说
，只要 emission 比允许的 wavelength 更长就都能通过。通俗的说就是 DsRed 的红
光 emission 可以通过为了蓝光或者绿光设立的 emission filter (如果是 long pass
而不是 band pass 的话），从而被检测到。如果不是有多个 color 看 correlation
的话，问题不大。 |
|
l**********1
发帖数: 5204 | 25 STORM living cell video recording barrier might be fixed by nonlinear
quantum
spectroscopy after 2017 AC (possible)
Alán Aspuru-Guzik Harvard Univ
his team one PNAS paper:
Quantum state and process tomography of energy transfer systems via
ultrafast spectroscopy.
Yuen-Zhou J et al. (2011)
PNAS 108:17615-17620.
key words:
processing ∣ open quantum systems ∣ quantum biology>
link:
//www.ncbi.nlm.nih.gov/pubmed/21997214
... 阅读全帖 |
|
l**********1
发帖数: 5204 | 26 STORM living cell video recording barrier might be fixed by nonlinear
quantum
spectroscopy after 2017 AC (possible)
Alán Aspuru-Guzik Harvard Univ
his team one PNAS paper:
Quantum state and process tomography of energy transfer systems via
ultrafast spectroscopy.
Yuen-Zhou J et al. (2011)
PNAS 108:17615-17620.
key words:
processing ∣ open quantum systems ∣ quantum biology>
link:
//www.ncbi.nlm.nih.gov/pubmed/21997214
... 阅读全帖 |
|
d****d
发帖数: 214 | 27 A conversation with Robert Lefkowitz, Joseph Goldstein, and Michael Brown
Ushma S. Neill and Howard A. Rockman
Published May 1, 2012
Today we shift the format of our Conversations with Giants in Medicine and
allow three of our most charismatic giants (Robert Lefkowitz, Joseph
Goldstein, and Michael Brown) to interview each other (Figure 1). Lefkowitz
(Duke University) is known for his seminal discoveries in understanding G
protein–coupled receptor function. The legendary partnership between Brow... 阅读全帖 |
|
|
b******y
发帖数: 627 | 29 As particles around a micron, these scatter light at 600nm, 500nm, or 400nm.
My guess bacteria don't have pigments that absorb at 600nm but does have
pigments that absorb at these lower wavelength.
Just my guess. |
|
o****e
发帖数: 1011 | 30 和Mg结合时有特定的、“long wavelength”的吸收时，研究起来才会容易些。
protein的“intrinsic fluorescence”(比如Trp)可影响的因素太多，我不会把
它作为首选的定量研究“Mg binding event”的手段。 |
|
m**z
发帖数: 787 | 31 fluoresent probe can be used for a competition experiment. Or use Co to
produce the long wavelength (d-d transition) and back titrate Mg. |
|
z*f
发帖数: 27 | 32 From Wiki:
Using the Beer-Lambert Law it is possible to relate the amount of light
absorbed to the concentration of the absorbing molecule. At a wavelength of
260 nm, the average extinction coefficient for double-stranded DNA is 0.020
(μg/ml)−1 cm−1, for single-stranded DNA it is 0.027 (μg/ml)&#
8722;1 cm−1, for single-stranded RNA it is 0.025 (μg/ml)−1 cm&#
8722;1 and for short single-stranded oligonucleotides it is dependent on the
length and base composition. Thus, an ... 阅读全帖 |
|
c*******0
发帖数: 190 | 33 http://www.nature.com/ncomms/2015/150915/ncomms9264/full/ncomms
Abstract
A major challenge in neuroscience is to reliably activate individual neurons
, particularly those in deeper brain regions. Current optogenetic approaches
require invasive surgical procedures to deliver light of specific
wavelengths to target cells to activate or silence them. Here, we
demonstrate the use of low-pressure ultrasound as a non-invasive trigger to
activate specific ultrasonically sensitized neurons in the nemato... 阅读全帖 |
|
T*****e
发帖数: 247 | 34 这没什么脸红的吧。你不要想它是重大意义，就是吹牛。你换个角度想，它的普遍意义
是什么。如果你要跟自己的家人讲自己的工作，你绝对不会从基因开始讲吧。跳出来看
看，做这些到底为了啥？
举个普遍意义但不很贴切的例子：
Do not start with, "I have been trying to explain the interesting wavelength
dependence of light scattering from small particles," but rather "There is
a widespread need to explain to one's kids why the sky is blue."
另一个角度是你的工作到底对这个领域有什么推进。而这个领域为什么存在，存在的意
义是什么，健康，疾病，而你的工作对认识这一切有了什么贡献？偷懒的方法就是找找
最近你们领域的综述，看看人家introduction和最后perspective对这个领域怎么总结
的。这就是重大意义了吧。 |
|
s*******1
发帖数: 188 | 35 非常好的建议，谢谢啊
wavelength
is |
|
s******r
发帖数: 1245 | 36 TALEN用双色的思路应该是发在cell还是csc上的，那是几年前了，所以idea本身不算是
很innovative。你在质粒上加个蓝色做负筛选，这个不知道有没有人做过，要没有的话
可以试试，如果你在那几个大牛实验室可能会发的还可以，不过看你在这儿问这些估计
不是，所以期望别太高。流式的时候不是几个颜色就照几次的，一道光照上去，收的时
候split，看光源用了几个wavelength。
所谓干细胞基因编辑效率低其实是个伪命题，如果你的目的是建系多用点细胞就好了，
基本上没有拿不到的。如果你是要研究提高效率方法，那么除非新办法是改进机制的能
实现数量级上的提升，其他的修修补补不过是小打小闹，就是为了灌一瓢水。
好像我都是在泼冷水，抱歉。 |
|
发帖数: 1 | 37 无意中看到一个老外写的，用了下面这篇文章里边介绍的使用荧光标签检控mitophagy。
Tools and techniques to measure mitophagy using fluorescence microscopy
DOI: 10.4161/auto.24001
FP-based biosensors for mitophagy
The field of FP engineering is accelerating with a plethora of new variants
described each year. These proteins show differen- tial properties, many of
which may provide useful tools to study mitophagy. For example, FP-based
biosensors such as a pH- sensitive mitochondrial-targeted FP chimera has
been recently developed to m... 阅读全帖 |
|
发帖数: 1 | 38 无意中看到一个老外写的，用了下面这篇文章里边介绍的使用荧光标签检控mitophagy。
Tools and techniques to measure mitophagy using fluorescence microscopy
DOI: 10.4161/auto.24001
FP-based biosensors for mitophagy
The field of FP engineering is accelerating with a plethora of new variants
described each year. These proteins show differen- tial properties, many of
which may provide useful tools to study mitophagy. For example, FP-based
biosensors such as a pH- sensitive mitochondrial-targeted FP chimera has
been recently developed to m... 阅读全帖 |
|
k***i
发帖数: 662 | 39 IN RELATED work, Heeger, Kwanghee Lee, and their coworkers demonstrated that tandem polymer solar cells—a design in which two cells are connected in series—can be fabricated entirely using inexpensive solution-phase methods. Connecting two cells made from semiconductors with distinct band gaps broadens the range of wavelengths the system can collect to generate electricity. In contrast to other researchers who have employed high-vacuum deposition methods to prepare tandem cells, the UC Santa Bar |
|
c*******e
发帖数: 170 | 40 From my experience, I dont think it is possible that the jet will induce the
dissociation of N2. You said there is resonant ionization by laser. I
would ask what kind of you are using, especially for the pulse duration and
wavelength.
Let's consider this problem in two ways:
1. Suppose what you have seen is really Nitrogen atomic ion. The most
likely mechanism should be Nitrogen molecule absorbs the photon(s) to
dissociation, and Nitrogen atom is ionized by other photons. However, the
highe |
|
j******0
发帖数: 258 | 41 Title: Multi-wavelength operation of optical disk drives for chemical
and biological analysis
Source: Sensors and actuators. B, Chemical
yr:2009
vol:136 iss:1
pg:203 -208 |
|
y****i
发帖数: 1504 | 42 shift your excitation wavelength to blue and/or
decrease excitation light intensity. |
|
t******t
发帖数: 3045 | 43 Just wanted to mention the relationship between line width and relaxation.
Line width is related to T2 while T2
is related to T1. If the relaxation in xy is slow, the relaxation in z is
also slow. The longer T2 is, the sharper
the peak is. The reason why NMR peaks are only a few Hz wide while UV-Vis
and IR peaks are much wider and
have to be measured by wavelength in nm is that the relaxations in these
spectroscopy are much faster.
The broadening caused by exchange is another story. If the excha |
|
j******0
发帖数: 258 | 44 Title: Multi-wavelength operation of optical disk drives for chemical
and biological analysis
Source: Sensors and actuators. B, Chemical [0925-4005] Potyrailo yr:2009
vol:136 iss:1 pg:203 -208 |
|
p*******g
发帖数: 13 | 45 How about using another wavelength which is more sensitive to your compound?
Or use another kind of detector? Like MS or CAD? |
|
x*******i
发帖数: 32 | 46 Hello there:
Trytophan fluorescence spectroscopy could be used to analyze whether protein
is compact or open. Ex at 288 nm and em around 320 nm suggests that protein
is compact (trytophan in hydrophobic environment). Em at longer wavelength
suggests an open structure. Also relative fluorescence intensity could tell
something? Anybody know why?
Thanks |
|
P***e
发帖数: 804 | 47 mobile phase B 10-30% gradient内 baseline linearly increase. 听起来像是
ACN的问题。Try higher wavelength such as 254 nm, if no baseline drifting,
most likely it is the ACN impuritiy problems.有没有试过不同batch或vendor 的
ACN？
题，但
30 |
|
A*******l
发帖数: 67 | 48 我们用Varian比较多
现在用的都是100和300系列
优点是stability, sensitivity,reproducibility
官方的baseline stability是0.0003 abs/hr,我实测到新的机器可以到0.0001abs/hr，
多次重复一致
缺点是维修比较贵，由于典型的double beam设计，旧仪器chopper元件的故障率很高，
基本上两三年要换一次，当然也是因为我们用的assay比较intense，经常7X20的用
一直很想买他家的50/60系列，但是老板不批钱
50/60系列的优点是速度和方便
像楼上讲的Agilent一样，可以开室测试。因为使用功率非常高的Xenon lamp，而不是
普通的UV/vis lamp，所以室内光源对测试基本没影响。而且由于Xenon lamp直接覆盖
190-1200nm，没有通常所见的uv/vis光源切换造成的spike。这个系列使用dual beam设
计，没有chopper元件，所以故障率比较低。缺点是理论上来讲没有double beam的
stability和reproducibility好，但实测几... 阅读全帖 |
|
A*******l
发帖数: 67 | 49 我觉得你说的monochromator是那种specific single wavelength light input device
.我见到的那台是single channel light emitter，连接别的detector 和sample
chamber用作fluorometer
很少有人把UV/vis叫monochromator吧，虽然原理上它包括某种monochromator作为光源。
Xenon flash lamp 和DT就看个人爱好。DT比较传统，但是除非你买5000/6000系列，不
然我觉得performance差不多。 |
|
s******g
发帖数: 177 | 50 Purity or potency?
If potency, standard is required and LC-MS works.
If purity, you can use peak area purity (low wavelength UV) |
|
