[gel-electrophoresis-data commit] r15 - trunk/lab-book-latex
codesite...@google.com
Date: Tue Jun 10 03:40:09 2008
New Revision: 15
Modified:
trunk/lab-book-latex/GibsonLabBook.dvi
trunk/lab-book-latex/GibsonLabBook.pdf
trunk/lab-book-latex/GibsonLabBook.tex
trunk/lab-book-latex/LabBook-abbrevs.tex
trunk/lab-book-latex/experiment.tex
trunk/lab-book-latex/protocol.tex
Log:
added experiment 03-18 Lowry protein quantification
Modified: trunk/lab-book-latex/GibsonLabBook.dvi
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Modified: trunk/lab-book-latex/GibsonLabBook.pdf
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Binary files. No diff available.
Modified: trunk/lab-book-latex/GibsonLabBook.tex
==============================================================================
--- trunk/lab-book-latex/GibsonLabBook.tex (original)
+++ trunk/lab-book-latex/GibsonLabBook.tex Tue Jun 10 03:40:09 2008
@@ -22,25 +22,25 @@
%new packages Frank has added
\usepackage[printonlyused]{acronym}
\usepackage{paralist}
-\usepackage[pdftex]{hyperref}
-\hypersetup{
- bookmarks=true, % show bookmarks bar?
- unicode=false, % non-Latin characters in Acrobat’s bookmarks
- pdftoolbar=true, % show Acrobat’s toolbar?
- pdfmenubar=true, % show Acrobat’s menu?
- pdffitwindow=true, % page fit to window when opened
- pdftitle={My title}, % title
- pdfauthor={Author}, % author
- pdfsubject={Subject}, % subject of the document
- pdfnewwindow=true, % links in new window
- pdfkeywords={keywords}, % list of keywords
- colorlinks=true, % false: boxed links; true: colored links
- linkcolor=blue, % color of internal links
- citecolor=green, % color of links to bibliography
- filecolor=magenta, % color of file links
- urlcolor=cyan % color of external links
- plainpages=false
-}
+%\usepackage[pdftex]{hyperref}
+% \hypersetup{
+% bookmarks=true, % show bookmarks bar?
+% unicode=false, % non-Latin characters in Acrobat’s bookmarks
+% pdftoolbar=true, % show Acrobat’s toolbar?
+% pdfmenubar=true, % show Acrobat’s menu?
+% pdffitwindow=true, % page fit to window when opened
+% pdftitle={My title}, % title
+% pdfauthor={Author}, % author
+% pdfsubject={Subject}, % subject of the document
+% pdfnewwindow=true, % links in new window
+% pdfkeywords={keywords}, % list of keywords
+% colorlinks=true, % false: boxed links; true: colored links
+% linkcolor=blue, % color of internal links
+% citecolor=green, % color of links to bibliography
+% filecolor=magenta, % color of file links
+% urlcolor=cyan % color of external links
+% plainpages=false
+% }
%\usepackage{attachfile}
Modified: trunk/lab-book-latex/LabBook-abbrevs.tex
==============================================================================
--- trunk/lab-book-latex/LabBook-abbrevs.tex (original)
+++ trunk/lab-book-latex/LabBook-abbrevs.tex Tue Jun 10 03:40:09 2008
@@ -32,6 +32,7 @@
\acro{CV}{Controled vocabulary}
\acro{CS}{New Born Calf Serum}
\acro{CY}{cytosol}
+\acro{DOC}{Deoxycholic acid}
\acro{DOLCE}{Descriptive Ontology for Linguistic and Cognitive Engineering}
\acro{DIGE}{difference gel electrophoresis}
Modified: trunk/lab-book-latex/experiment.tex
==============================================================================
--- trunk/lab-book-latex/experiment.tex (original)
+++ trunk/lab-book-latex/experiment.tex Tue Jun 10 03:40:09 2008
@@ -521,7 +521,67 @@
\end{center}
\end{table}
+\section{03-18: Lowry Procedure Using Samples from 03-07}
+\subsection{Protocol}
+Seven aliquots from Day 12 of Experiment 03-07, (Section \ref{sec:03-07}) were defrosted and pooled together. 500 $\mu$l were used in this protein assay, (See Protocol \ref{sec:lowry}) the rest were frozen down in 1 ml aliquots.
+
+\subsection{Results}
+
+The protein standards were made up as in Table \ref{tab:protein-stand}. The volume of buffer was added so to match the concentration of the cells in the sample. Each experiment was repeated twice.
+
+
+\begin{table} [ht]
+\caption[Lowry Protein concentration]{Lowry protein concentration}
+\begin{center}
+
+\begin{tabular}{| p{3cm} |p{3cm} |p{3cm} |p{3cm} | p{3cm} }
+\hline
+ Tube & Volume of stock ($\mu$l) & H20 ($\mu$l) & Buffer ($\mu$l) & Protein Concentration ($\mu$g/ml)\\
+\hline
+ 1 & 0 & 850 & 150 & 0 \\
+ 2 & 25 & 825 & 150 & 10\\
+ 3 & 50 & 800 & 150 & 20 \\
+ 4 & 100 & 750 & 150 & 40 \\
+ 5 & 150 & 700 & 150 & 60 \\
+ 6 & 200 & 650 & 150 & 80 \\
+ 7 & 250 & 600 & 150 & 100 \\
+ 8 & 375 & 475 & 150 & 150 \\
+ 9 & 500 & 350 & 150 & 200\\
+\hline
+\end{tabular}
+\end{center}
+\label{tab:protein-stand}
+\end{table}
+
+
+Samples were made up as in Table \ref{tab:protein-stand-sample} and the experiment was repeated 3 times.
+Assuming the concentration of protein in the cell sample is 6$\mu$g/ml therefore
+
+
+\begin{itemize}
+ \item 10 $\mu$g/ml = 1.5 $\mu$l
+\item 100 $\mu$g/ml = 15 $\mu$l
+\item 1000 $\mu$g/ml = 150 $\mu$l
+\end{itemize}
+
+\begin{table} [ht]
+\caption[Lowry Protein concentration for samples]{Lowry protein concentration for samples}
+\begin{center}
+
+\begin{tabular}{| p{3cm} |p{3cm} |p{3cm} |p{3cm} | p{3cm} }
+\hline
+ Tube & Sample ($\mu$l) & Buffer ($\mu$l) & H20 ($\mu$l) \\
+\hline
+ A & 1.5 & 148.5 & 850 \\
+ B & 15 & 135 & 850 \\
+ C & 150 & 0 & 850 \\
+
+\hline
+\end{tabular}
+\end{center}
+\label{tab:protein-stand-sample}
+\end{table}
Modified: trunk/lab-book-latex/protocol.tex
==============================================================================
--- trunk/lab-book-latex/protocol.tex (original)
+++ trunk/lab-book-latex/protocol.tex Tue Jun 10 03:40:09 2008
@@ -258,8 +258,63 @@
% \begin{figure}
% \centering
-% \includegraphics{lab-book-images/SubcellularFractionation.png}
+% \includegraphics[scale=0.5]{lab-book-images/SubcellularFractionation.png}
% % SubcellularFractionation.png: -1238670960x-1238695792 pixel, 0dpi, 1232004255318016.00x1232028951379968.00 cm, bb=
% \caption{Subcellular Fractionation}
% \label{fig:subcellular-fractionation}
% \end{figure}
+
+
+\section{Protein quantification by Lowry Procedure}
+\label{sec:lowry}
+
+\begin{enumerate}
+ \item In the appropriately labelled plastic microcentrifuge tubes, dilute the stock protein standard solution in a total volume of 1 ml. The standard solutions are shown below in Table
+
+
+\begin{table} [ht]
+\caption[Lowry Protein Standards]{Lowry Protein Standards}
+\begin{center}
+
+\begin{tabular}{| p{3cm} |p{3cm} |p{3cm} |p{3cm} |}
+\hline
+ Volume of 400 $\mu$g/ml protein standard solution ($\mu$l) & Volume of dieonised water ($\mu$l) & Protein concentration ($\mu$g/ml) \\
+\hline
+ 0 & 1000 & 0 \\
+ 25 & 975 & 10 \\
+ 50 & 950 & 20 \\
+ 100 & 910 & 40 \\
+ 150 & 850 & 60 \\
+ 200 & 800 & 80 \\
+ 250 & 750 & 100 \\
+ 375 & 625 & 150 \\
+ 500 & 500 & 200 \\
+\hline
+\end{tabular}
+\end{center}
+\end{table}
+
+
+\item Label the microcentrifuge tube containing the protein concentration 0$\mu$g/ml as the ‘BLANK’.
+\item Put the sample(s) in microcentrifuge tube(s) and label. Again, dilute to a total volume of 1 ml
+\item Add 100$\mu$l of the \ac{DOC} solution to each of the three types of tube (STD, BLANK and SAMPLE)
+\item Vortex mix then leave for 10 minutes at room temperature.
+\item Add 100$\mu$l \ac{TCA} to all tubes and vortex mix.
+\item Pellet the precipitates by centrifugation for 10 minutes at max revs.
+\item Decant and blot away the supernatants.
+\item Add 1 ml of Lowry modified reagent to each tube then allow the pellets to dissolve.
+\item Transfer the solutions to their correspondingly labelled test tubes.
+\item Rinse each microcentrifuge tube with 1ml of water and add the rinsings to their respective test tubes.
+\item Ensure the contents of the tubes are mixed well and leave the solutions to stand for 20 minutes at room temperature.
+\item With rapid and immediate mixing, add 0.5ml Folin and Ciocalteu’s Phenol reagent to each tube.
+\item Leave tubes to stand at room temperature for 30 minutes. Purple/blue colour should now develop.
+\item At this stage turn on spectrophotometer, to allow it to warm up.
+ \item Transfer the solutions to cuvettes for measurement of absorbence at a wavelength between 500nm and 800nm (usually 700nm).
+\item Measure the absorbence of the STD and SAMPLES against the BLANK. Readings should be completed within 30 minutes.
+\item Create a calibration curve by plotting the absorbence of the STDs against their known concentrations.
+\item From this calibration curve calculate the concentration of protein in the SAMPLE tube(s). This is done by multiplying the results by the dilution factor to obtain the protein concentration in the original sample.
+\end{enumerate}
+
+
+
+