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From CellBiology

--Z3272558 15:13, 8 March 2012 (EST)

Lab Attendance

Lab 1

--Z3272558 15:37, 8 March 2012 (EST)

Lab 2

--Z3272558 14:59, 15 March 2012 (EST)

Lab 3

--Z3272558 14:25, 22 March 2012 (EST)

Lab 5

--Z3272558 14:17, 19 April 2012 (EST)

Lab 8

--Z3272558 14:20, 3 May 2012 (EST)

Lab 11

--Z3272558 14:17, 17 May 2012 (EST)

Lab 12

--Z3272558 15:39, 31 May 2012 (EST)

Notes

Lab 1

No notes.

Lab 2

Different microscopic techniques and their applications in cell study.

Lab 3

Preparation and Fixation

Aims of the process:

1. Preserve cell structure - prevent autolysis

2. Inhibits bacterial and fungal growth (preserves)

3. Make the tissue resistant to damage during subsequent processing (hardy)- robust

4. Allow access of stains and antibodies (permeable)- done by fixation

Safety Data Sheet (SDS)

Universal labelling and identification standard on chemicals

3 main techniques for fixation

1.Fresh Frozen - rapidly freezing tissue

2.Precipitation - Immersion in cooled organic solvents- methanol or acetone or acids

3.Aldehyde Cross-linked - Cross-links are generated between several reactive groups (mainly -NH2 groups) such as found in protein lysine residues.


Lab 4

Immunohistochemistry

Introduction to immunological methods for analysis of cells and tissues in cell biology.

Immunochemistry can also be called immunohistochemistry or immunocytochemistry.

Lab 4 Exercise

Musashi (Msi) is an evolutionarily conserved gene family of RNA-binding proteins (RBPs) that is preferentially expressed in the nervous system. It is a stem cell marker. A type of RNA-binding protein. 35 kDA mass.

Shinsuke Shibata, Masahiko Umei, Hironori Kawahara, Masato Yano, Shinji Makino, Hideyuki Okano Characterization of the RNA-binding protein Musashi1 in zebrafish. Brain Res.: 2012, 1462;162-73 PubMed 22429745


Primary antibody : Rabbit IgG

It is a type of polyclonal antibody and can be found on rabbits, goats, chicken and mouse. IgG.

Musashi-1

Musashi-2


Secondary antibody : Anti-Rabbit IgG antibody [LO-RG-1] - Rat Monoclonal

This product should be stored undiluted. Avoid repeated freezing and thawing as this may denature the antibody.

Dilution : 500 µg at 1mg/ml

Isotype : IgG2a

Anti Rabbit IgG

Lab Assessment 2 : Microscopy

Cell Leakage.jpg

G MAJNO, G E PALADE Studies on inflammation. 1. The effect of histamine and serotonin on vascular permeability: an electron microscopic study. J Biophys Biochem Cytol: 1961, 11;571-605 PubMed 14468626



Rockefeller University Press Copyright Policy This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.jcb.org/misc/terms.shtml). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).


PMID:

   14468626


Lab Assessment 3 : Fixation

Locate a current SDS for one of the fixatives described in today's lab. Identify the properties and hazards associated with that chemical.

SDS : Paraformaldehyde

Paraformaldehyde SDS


Properties of Paraformaldehyde

1. White solid, formaldehyde-like odor

2. pH : neutral

3. Stability : Slowly sublimes to Formaldehyde

4. Solubility : Soluble in hot water (not soluble in Ethanol)

5. Odor : Pungent


Hazards associated with Paraformaldehyde

1. Eye : May result in corneal injury. Causes severe eye irritation and burns and also conjunctivitis.

2. Skin : May cause allergic reaction of the skin.

3. Ingestion : Harmful if swallowed. May cause severe digestive tract irritation with abdominal pain, nausea, vomiting and diarrhea.

4. Inhalation : May cause severe irritation of the upper respiratory tract with pain, burns, and inflammation. May also cause acute bronchitis.



Lab Assessment 4 Exercise

Musashi (Msi) is an evolutionarily conserved gene family of RNA-binding proteins (RBPs) that is preferentially expressed in the nervous system. It is a stem cell marker. A type of RNA-binding protein. 35 kDA mass.

Shinsuke Shibata, Masahiko Umei, Hironori Kawahara, Masato Yano, Shinji Makino, Hideyuki Okano Characterization of the RNA-binding protein Musashi1 in zebrafish. Brain Res.: 2012, 1462;162-73 PubMed 22429745


Primary antibody : Rabbit IgG

It is a type of polyclonal antibody and can be found on rabbits, goats, chicken and mouse. IgG.

Musashi-1

Musashi-2

Lab Assessment 6 : Chart

Phenotype Chart Group 3.JPG

Group 3 - Analysis of morphological phenotypes in Tropomyosin 4 over-expressing B35 neuroepithelial cells

Group A is TM4 and Group B is Control

Lab Assessment 7 : Group project contribution

Task divided and we have put up some content into the page. Introduction and history is done and we are now working on the other parts of the presentation.


Lab 8 : Tissue Culture

Identify a mammalian cell line in the ATCC catalogue (and add a link)

NCTC clone 929 [L cell, L-929, derivative of Strain L]

ATCC Cell Line

Identify the original tissue of origin of that cell line.

Organism: Mus musculus Morphology: fibroblast Tissue: subcutaneous connective tissue; areolar and adipose Strain: C3H/An

Identify the original paper that characterised the properties of that cell line.

M H Hutz, A M Michelson, S E Antonarakis, S H Orkin, H H Kazazian Restriction site polymorphism in the phosphoglycerate kinase gene on the X chromosome. Hum. Genet.: 1984, 66(2-3);217-9 PubMed 6325324


Lab Assessment 9 : Peer Review

Peer Assessment of Group Projects

Group 1

The introduction is short and provides a good brief summary of the topic but it needs more information. Well organised, interesting use of the tables. But too much use of the tables make it look cluttered at some times. All of the parts were almost complete and done, which is good and showed that the group is functioning very well. Addition of a student drawn diagram would make the page a bit more interesting as well. Some images in the clinical uses of testosterone might make the page less wordy and more interesting. I would say the use of dot points would make the page more user-friendly.

Group 2

The image at the top of page should have caption so people know what they are looking at. Intro and history are good, maybe some more information can be added to intro so that it becomes more interesting. The tables are dull maybe because of its colour scheme? Some more info on the pathway would be good to the readers. The abnormal function is very engaging and very easily understood particularly because of the layout of the table.

Group 3

Introduction and history looks good by none of the information were cited. In my opinion, the history would look a lot more better being put in a table. Signalling pathway is again not cited and the use of diagrams would make it easier for people to understand, particularly because this topic is not very common to the public. The use of flow diagram is good for this. Obviously the page is still not done with none of the glossary being done and most no citation being shown on the page.

Group 4

Good introduction and a very well presented history of Notch pathway using the table. The signalling pathway can be aided by the use of diagrams instead of just merely words. It appears that it's very short in my opinion. The normal function bit was very nicely done, but other than that there is not headings for abnormal pathway. Clinical use section would make it more interesting as people can understand the use of this pathway in real life.

Group 5

Introduction and history was very good. The layout was very good and consistent going form each subheadings and everything seems to be cohesive. There is continuity in each section making it very intriguing for readers. All key points relating to the topic are present. The project is full of proof that the topic has been researched to a suitable depth and it contains and element of teaching and this is shown through dot points, text, diagrams, images and videos. The referencing and glossaries were nicely done and this project is very effectively done between the group members.

Group 6

Intro, history and structure of insulin is shown very clearly. The introduction has quite a bit of information to it. The structure of insulin: the picture is a good attribute but i do think a bit more insulin itself might help here. The signalling pathway is well-covered and the picture complements your text well. Normal function is better with more citations.There is quite a lot of information in abnormal function as well, which demonstrates more effort has been put into this section. A summarizing table would be good for this. Interesting to read what is currently being researched. Few more terms to your glossary would make it better. The references seem to be fine and well done as a whole.

Group 7

The page is very well planned and researched. It seems like all the group members have put effort into it. The balance between pictures and text is very good and the intro could be a bit shorter perhaps. The self drawn diagram is very good. All the pictures were cited correct and accordingly. For all the information provided the journal articles used are very less for e.g. the 1st paragraph of inhibitory pathway has no citation at all and the regulation of beta arrestin has one article ref. Maybe put more citation so that it shows a lot of research has been done prior to writing? The normal/abnormal function was very good and detailed.


--Mark Hill 13:46, 17 May 2012 (EST) You have not completed the peer assessment process yet. If you have made comments on each project page they need also to be pasted here today for me to include in your individual assessment.

Lab 12: Microarray

Identify a current technique used in gene sequencing.

Nanopore technology is used to sequence genes and it was developed by Oxford University. Various companies are licensed to use the technique nowadays.The end-to-end system includes sample preparation, molecular analysis and informatics, and is designed to provide disruptive user benefits in a number of applications. The techniques involves nanopore fabrication and nanopore sensing.

Identify a recent cell biology research paper that has used microarray technology.

The paper titled "Isolation of high quality RNA from embryonic kidney and cells." is one of the recent research in the cell biology field that uses the microarray technology. In this study, methods for the identification of the specific mRNAs and the quantitation of their levels are used in understanding gene expression and one of the main tools used was the microarray technique.

Shifaan Thowfeequ, Odyssé Michos Isolation of high quality RNA from embryonic kidney and cells. Methods Mol. Biol.: 2012, 886;203-10 PubMed 22639263


What aspect of the research findings were contributed by the microarray technique.

The methodology for isolating high quality RNA from embryonic kidneys for various applications include microarray analysis and quantitative reverse transcription PCR (qRT-PCR).This is where microarray plays its role in the research.

Links

http://www.wikipedia.com

LECTURE 2