Easy data template, analysis and presentation tools for complex ...

Volume 4, Issue 1, 2011
Easy data template, analysis and presentation tools for complex multiparametric animal
cell culture studies
Maddaly Ravi, Associate Professor, Sri Ramachandra University, maddalyravi@hotmail.com
Sai Keerthana W, UG student, Sri Ramachandra University, keerthanaw@gmail.com
Madhumitha Haridoss, PG student,Sri Ramachandra University, h.madhumitha@gmail.com
Shwethaa Raghunathan, UG student,Sri Ramachandra University, vrisha.rox.da.wrld@gmail.com
Sowmya Thanikachalam, PG student,Sri Ramachandra University, saumyasubramanium@gmail.com
Abstract
Cell culture experiments are important for biological education and research. Standardization of optimal
conditions to obtain 3 Dimensional aggregates of a cell line was performed which involved studying the
effects of several parameters with various combinations of the variables. Data records, analysis and
interpretations become voluminous and complex with increasing variables. To simplify and semi automate
this process, an excel sheet and a single table format was designed to represent the results of various
combinations of the variables studied. These formats can be effectively used for science education and
research as a methodology by all working with cell cultures.
Introduction
Animal cell cultures have become indispensable for a variety of uses and are integral to biological education
globally. Cell cultures and experiments involving them are important components of undergraduate,
graduate and research programmes in all fields of biological sciences. The reason is that once the required
skills are acquired, cells in culture are reliable models for scientific studies. The culture techniques have
evolved with simultaneous developments in supportive requirements for cultures such as media and
supplements. [1] Cultured cells are being widely utilized for many studies including cell physiology,
molecular biology and cancer research to mention a few. [2] Cell cultures are crucial for drug discovery and
validation studies. The revolutionary Hybridoma technology to produce monoclonal antibodies is essentially
a conglomeration of different cell culture methodologies. From cultures as 2 Dimensional models, the
methodology has developed into culturing cells in a 3 Dimensional environment. These models are gaining
importance in areas such as cancer research, tissue engineering etc., through better understanding of gene
expressions and protein profile changes. From cells in cultures being the base for many products, they have
now themselves become the products for direct applications.
Cells can be cultured in variety of culture vessels that include flasks, plates and petri dishes. The choice of
the culture vessel depends on the number of cells required, type of study, number of variables being tested
and also the number of study point ranges within each of the study variable. When large cell numbers are
required, culture flaks are convenient. When many variables are used in the study, such as the effects of
more than one drug or various concentrations of one or more drugs, it is easier to work in culture plates. The
6, 12, 24 and 96 well plates where studies involving multiple variables can be conducted conveniently.
For documentation, representation and analysis, it is imperative to have study design representative
templates such as in the form of Tables in the word file, etc. The level of difficulty in such a word file, as a
Table becomes increasingly difficult with increasing number of variables used in a study. For example, it can
be tedious to type all the variables and parameters in each of the well area for a 24 well plate in a Table
form of a word document. Multi well plates are also used to maintain cell cultures in different conditions.
Parameters such as media used, seeding density, volume of culture etc. are varied for different cell lines for
every culture that is set up. Creation of the tables and data entry to record the information for plate
templates separately is time consuming. The software programme Excel has been used for various
applications in the field of biology from analysis of statistical data to maintenance of lab records. [3] MS
Excel was used for automating data entry for varied parameters used in animal cell culture and creating
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templates for 6-well, 12-well and 24-well cell culture plates. To automate and facilitate multi parametric
analysis of data, a tool to tabulate diverse components involved in cell culture on a single platform was
adopted as previously described. [4] The data was entered into an excel file to create a plate template.
Programming the cells on the plate template was made such that the data entered in previously is
automatically displayed in the plate template. Likewise, once the results are obtained, the need to represent
the net result of a complex study in a single format is very useful for several reasons including significant
reduction in the numbers of entities such as Tables/Graphs, etc and also for easier comparisons of the
multiple variables. A single table where different parameters can be analyzed and represented was
developed towards this. These innovative tools have applications for easy data presentation, analysis and
interpretation and also as important components of teaching, learning and research at the undergraduate
and graduate level practical sessions involving cell cultures.
Methodology
Cell culture plate layout designs with experimental multiparametric details as an excel file
The parameters studied
The parameters considered for the study include well identification number, volume of medium added for
plating, concentration of agarose in the medium, the medium in which agarose was dissolved, presence or
absence of conditioned medium and the cell seeding density.
Data entry
An excel sheet was opened and different columns were named according to the different parameters
involved in various wells of a cell culture plate as represented in Figures 1 and 2.
Creation of a template
A template for 6 well, 12 well and 24 well plate was created using “merge and centre” for the selected cells.
The border for the template was given in the following series of steps:
The entire template was selected
The selected template was left clicked on
“Format Cells” was selected
The tab “Border” in the dialogue box was chosen
The options “Outline” and “Inside” and click on “OK” were selected
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Figure 1. Data entry into excel cells for the fields well identity, agarose volume, % agarose composition,
medium used for dissolving agarose, presence or absence of conditioned medium and seeding densities.
Programming the excel sheet cells
The cells corresponding to the culture plate wells were programmed to input data from the excel sheet using
‘CONCATENATE’ function to merge data from the previously entered information at the top of the excel
sheet. The programming was done by the following series of steps and represented in Figure 3.
The cells to be programmed were selected
The formula tab was clicked and concatenate formula was selected.
It is a text function used to join several text strings into one text string and parenthesis is used to enter
the formula for programming the cell. The formula starts with a double quotes and ends with a double
quote:
=CONCATENATE("",D2," of ",E2," % Agarose in ",F2," with a SD of ", H5," M (CM",G2,")")
Figure 2. The template has been created using ‘Merge and Centre’ Function and choosing
‘Wrap Text’ option and the borders are framed using ‘Format Cells’
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Figure 3. Programming of the Cells using ‘CONCATENATE’ function to merge many text strings from different
cells into single output cell.
Conditional Formatting of the cells
Conditional Formatting offers and easy way to apply formats that stay dormant until the values in the highlighted
cells achieve a specified state as given in Figure 4. Here a tool was used to highlight cells with a certain color
depending on cell value. The series of steps are as follows:
The cells to be highlighted depending upon the condition were selected
The option “Conditional Formatting” and select “New Rule” was clicked on
The second option “Format only cells that contain” was selected and “specific text” in the dropdown box
under “Edit the rule description” was subsequently selected and the value was entered.
“Format” was clicked and different options such as number, font, border, fill etc., were used
“Ok” was clicked to allow conditional formatting.
On conditional formatting, different parameters are represented in different ways such as:
The different cells containing PBS and RPMI are shown in different shades
Cells with conditioned medium are shown by double underlining
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Figure 4. Conditional Formatting of the programmed cells was used to change parameters in the format of the
template to distinguish between different conditions in the cell culture plate
Representation of analysed data
In studies that involve series of experiments to standardize the 3D cell culture conditions, various variables
and combinations of the variables have to be observed, data recorded and analyzed. For this purpose, a
single template to analyze and represent all the parameters to record the results was developed. The
parameters include concentration of agarose, culture surface area, types of culture medium, culture medium
volumes, presence or absence of conditioned medium and various seeding densities. Thus the result
entered in a single box is a representative of a combination of all these parameters as shown in Table 1.
CONCENTRATION OF AGAROSE
CM 0.25% 0.5% 0.75%
+/- 6 12 24 6 12 24 6 12 24
PBS
+ 1
ml
-
+ 750
µl
-
+ 500
µl
-
+ 250
µl
-
V
O
L
U
M
E
+ 1
ml
-
RPMI
+ 750
µl
-
+ 500
µl
-
+ 250
µl
-
+/- X Y X Y X Y X Y X Y X Y X Y X Y X Y
SEEDING DENSITIES
Table 1. The template provides for various parameters including concentration of agarose, medium of cell
suspension, volume of medium added, presence or absence of conditioned medium (CM) and seeding
densities per well in a 6 well, 12 well & 24 well plate.
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Results
An excel file was developed in a series of steps with simple formulae to make data entry and record keeping
simpler for multiparametric cell culture studies. Also, a single table format was developed which can serve
as a template for simultaneous data entry, result representation and analysis of multiple parameters involved
in cell culture studies.
Discussion
Cell cultures offer with excellent in vitro experimental models and should be well standardized for useful
study results. [5] Standardization of the optimal cell types and the culture types/conditions require multiple
experimental variables. Data analysis and interpretation is the most important and critical part of education
and research. The raw data obtained from experiments should be presented in an organized manner so that
useful information can be extracted from it. The excel template we describe here serves the required
purpose of easy data entry along with maximum information as obtained in the final template. Also, the
results obtained from all the variables/combinations of the variables could be entered into the single table
format that we describe here. The advantages of these two tools (the excel template and the single table
format) is requirement of less time, less confusion, less error and inclusion of maximum data. In addition, the
formats are flexible to suit a variety of experimental circumstances. These inputs are designed to refine and
distill the data so that students and researchers can glean interesting/pertinent information without the need
to sort through voluminous data or a number of single entities such as graphs and tables.
Conclusion
Science education is a combination of imparting information, instilling the need for an analysis, asking the
right questions and to choose the right approach thus contributing towards knowledge enhancement. In all
areas of life sciences, basic research is a vital part of science education. Cell cultures owing to their
immense potentials for both teaching and learning have become important components in science curricula
globally. Data analysis and interpretation is the critical part of any research which involves multiple
parameters and numerous variables. The cell culture plate template designed using MS Excel and table
generated using MS Word that are described here present dense data in a simplified form. The generated
data could be inspected, transformed and modeled with the useful and most pertinent information
highlighted. These are intended to considerably reduce the complexity of data record keeping, presentation
of the study design, analysis of results and sharing of the research findings. These tools can be very useful
for both learning and teaching for practical education in biological sciences.
References:
[1] R. I. Freshney, A. R. Liss (1994) Culture of Animal Cells: A Manual of Basic Technique, 3rd ed., New
York, Blackwell Publishing.
[2] Adi F. Gazdar, Girard Luc, Lockwood William W., Wan L. Lam and John D. Minna (2010) Lung cancer
cell lines as tools for biomedical discovery and research, J. Nat. Cancer Inst. 102, 1310–1321.
[3] D. Z. Meyer, L. M. Avery (2009) Excel as a qualitative data analysis tool, Field Method. 21, 91–112.
[4] M W. Lehman, C E. Herring (2003) Creating interactive spreadsheets to provide immediate feedback, J.
Account. Educ. 21, 327–337.
[5] J. H. Dunham, P. Guthmiller (2008). Doing good science: authenticating cell line identity, Cell Notice. 22,
15–17.
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