ethics to collect human biopsy and disposal

1. Guided by
Dr. Venu Talla
OVERVIEW ON HUMAN
CANCER CELL LINES
Presented by
J. Vishnu
RT/2016/604

2. CONTENTS
Introduction
History
Types of cell lines
Human cancer cell lines
Types of contamination
Disposal procedure
Advantages & Disadvantages
Summary
References
Overview on human cancer cell lines2

3. Introduction
 Definition:-
 Cell culture is the process by which cells are grown under controlled conditions,
generally outside of their natural environment.
 What is cell ?
 A cell is the smallest unit of life that can replicate independently, and cells are
often called the "building blocks of life". The study of cells is called cell biology.
 What is primary cell line ?
 Freshly isolated cells in culture until the first passage into a subculture
Overview on human cancer cell lines3
Dieter F, Methods In Animal Cell Culture, Institute of Biology, Department of Biophysics, University of Stuttgart

4.  What is Cell Line?
 After the first subculture, the primary culture becomes known as a cell line or
sub-clone.
 What is Cell Strain ?
 If a subpopulation of a cell line is positively selected from the culture by cloning
or some other method, this cell line becomes a cell strain
Overview on human cancer cell lines4
Cont…
Dieter F, Methods In Animal Cell Culture, Institute of Biology, Department of Biophysics, University of Stuttgart

5. Milestones
Overview on human cancer cell lines5
 Roux 1885: Embryonic chick cells were maintained in saline
 Harrison 1907: Amphibian spinal cord cultivated in lymph produced axons in vitro
 Carrel 1913: Cells in a culture can grow continuously under aseptic conditions
 Earle 1943: First continous cell line from L-cell mouse fibroblast.
 George Gey et al.,1952: Established continuous cell lines derived from human
cervical carcinoma – HeLa cells
 Eagle 1955:Developed by a systematic study a Basal medium of the nutrients and
the serum for cell culture
 Hayflick and Moorehead 1961: Human fibroblasts in culture have finite life time

6. Types of Cell Lines
Overview on human cancer cell lines6
Finite Cell Lines
•The cell lines with limited culture life
spans are referred to as finite cell
lines.
•The cells normally divide 20 to 100
times, the actual number of
doublings depends on the species,
cell lineage differences, culture
conditions etc.
•The human cells generally divide
50-100 times, while murine cells
divide 30-50 times before dying
Continuous Cell Lines
•Some cell lines become immortal
through a process called
transformation.
•Which can occur spontaneously or
can be chemically or virally induced.
•When a finite cell line undergoes
transformation and acquires the
ability to divide indefinitely, it
becomes a continuous cell line.
C.P. Kashyap et al, Human cancer cell lines- A brief communication, Journal of Chemical and Pharmaceutical Research; 2011, 3(6):514-520

7. Overview on human cancer cell lines7
Cell lines can be
propagated as two types
of cultures
Suspension culture.
Anchorage
independent
Cell cultures from
hematopoietic cells.
Ex: Leukemia cells,
Multiple melanoma cells.
Cell cultures from organ or
tissues.
Ex: Epithelial cells,
Fibroblasts.
Monolayer culture.
Anchorage dependent

8. Preparation of primary cell lines
Overview on human cancer cell lines8

9. Overview on human cancer cell lines9
Guidelines to collect the cells form human
 There are no guidelines or legislations regarding the use of human tissue in
research and in the biomedical industry in India
 The Indian Council of Medical Research (ICMR) guidelines, 2000, broadly cover
the categories of evaluation of drugs and diagnostics, epidemiological research,
human genetics research, transplantation research and assisted reproductive
technology
 In UK two acts are there
 Human Tissue (Scotland) Act 2006 applies only to tissue from the dead
 Human Tissue Act 2004 encompasses tissue both from the dead and living
Use of human tissues for research: Ethical concerns, Indian Journal of Medical Ethics Vol :3 :2008

10. Ethics to collect the human tissue
Overview on human cancer cell lines10
 Patient consent should only be taken by suitably qualified individuals with the
required specialist training
 The Patient Consent Form and associated Patient Information Sheet should be
written in concise and easy language that anyone can easily understand
 Explaining clearly the need for the specimen, the overall objective of the research
and why it is important (in lay terms)
 The additional discomfort or inconvenience that will occur if the donor agrees to
the request should be clearly explained
 The donor should be told clearly that there is no obligation whatsoever to
participate in the research
 If the research may be exploited commercially, the donor should be told clearly
what financial benefit might be gained from the research and a waiver to
commercial rights should be requested.
 R J Geraghty et al,Guidelines for the use of cell lines in biomedical research, British Journal of Cancer (2014) 111,1021–1046

11. Cont…
Overview on human cancer cell lines11
 All forms should be marked Confidential
 The information sheet and consent form must be printed on official-headed
notepaper.
Consent forms should address the following questions:
1. Have you read the information sheet about this study?
2. Have you had an opportunity to ask questions and discuss the study?
3. Have you received satisfactory answers to all your questions?
4. Have you received enough information about this study?
5. Which doctor have you spoken to about this study?
6. Do you understand that you are free to withdraw from this study (i) at any time, (ii)
without giving a reason and (iii) without affecting your future medical care?

12. Human cancer cell lines
Overview on human cancer cell lines12
 Cancers are a large family of diseases that involve abnormal cell growth with the
potential to invade or spread to other parts of the body by forming a subset
of neoplasm.
 Neoplasm or tumor is a group of cells that have undergone unregulated growth
and will often form a mass or lump, but may be distributed diffusely
Types of cancer
Colorectal cancer
Pancreatic cancer
Esophageal cancer
Prostate cancer
Kidney cancer
Skin cancer
Leukemia
Thyroid cancer
Bladder cancer
Lung cancer
Brain cancer
Melanoma
Breast cancer
Cervical cancer
Ovarian cancer
Liver cancer

13. Human cancer cell lines
Overview on human cancer cell lines13
Cell line Tissue &
organ
Age &
ethnicity
Morphology Disease
HT-1376 Human
bladder
58 (Caucasian-
F)
Epithelial Grade- III Carcinoma
HL-60 peripheral
blood
36(Caucasian-
F)
Myeloblastic acute pro-myelocytic
leukemia
MCF-7
Human breast 69(Caucasian-
F)
Epithelial Adenocarcinoma
SW 626 Ovary 46(Caucasian-
F)
Epithelial Grade-III
Adenocarcinoma
A172 Brain 53 ( Caucasian
-M)
Fibroblast Glioblastoma
MES-SA Uterus 56(Caucasian-
F)
epithelial Uterine sarcoma
NCI-H1573 Lung 35(Caucasian-
F)
Epithelial Adenocarcinoma stage-
IV
HELA Human, cervix 31( black-F) Epithelial Adenocarcinoma
HT-29 Colon 44(Caucasian- Epithelial Colorectal

14. Overview on human cancer cell lines14
 Glyn, Stacey the text book of cancer cell culture Humana Press vol.731: 79-91:2011
Types of
contamination
Biological
contamination
Easley detectable
Ex:-bacteria, molds,
yeast
Difficulty to detect
Ex:- viruses,
protozoa,
mycoplasma
Chemical
contamination
Presence of Non
living substance
ex:-media, serum,
Endotoxins,
Fluorescent

15. Disposal procedure
Overview on human cancer cell lines15
• Liquid disposal:-
1. For each 100 milliliters of blood or body fluid and cell lines, use 10 milliliters of
sodium hypochlorite (bleach)
2. Mix and let stand 30 minutes
3. Pour the mixture down a lab sink followed by a ten-fold excess of water
4. Alternatively, liquid waste may be autoclaved for 30 minutes at 121°C and
15psi
• Solid disposal
 Solid biological waste, e.g., pipettes, tissue culture flasks, and multiple well
plates, is typically deactivated by autoclaving
 Autoclave for 30 minutes at 121°C and 15psi

16. Needles, Syringes, Glass pipettes
Overview on human cancer cell lines16
 Autoclave your sharps container for a minimum
of 30 minutes at 121°C and 15psi
 Place sharps in a container that is red, rigid,
puncture resistant, leak-proof and labeled with
the biohazard symbol
 Log the autoclave run duration, quantity of
processed waste, date, and operator
 Label the sharps container with the words
“autoclaved”

17. Disadvantage
Overview on human cancer cell lines17
Advantage
Daniela Ferreira et al, The Importance of Cancer Cell Lines as in vitro Models in Cancer,1-22:2012
 Easy to handle and manipulate
 High degree of similarity with the
initial tumor
 High variety available
 Immediate accessibility
 Unlimited auto-replicative source
 Easy substitution
 Reproducibility of results
 Cross contamination with Hela cells
 Loss of heterogeneity
 Genomic instability
 Possibility of modifying the
characteristics of the cells
 Infections with mycoplasma
 Difficulty in the establishment of
long-term cancer cell lines
 Different environment of the tumor

18. Summary
Overview on human cancer cell lines18
 Human cell cultures will provide the tools for tissue engineering, gene therapy and
the understanding of protein function
 Human cancer-derived cell lines are the most widely used models to study the
biology of cancer and to test hypotheses to improve the efficacy of cancer
treatment
 The major problem with human cell lines is contamination
 By maintaining sterile condition we can prevent the contamination to the cell lines

19. Overview on human cancer cell lines19
References
 American Type Culture Collection (www.atcc.org)
 National Centre for Cell Science (www.nccs.res.in)
 European Collection of Animal Cell Cultures (ECACC)
 Biosafety in Microbiological and Biomedical Laboratories, 5Th Edition at
www.cdc.gov/od/ohs/biosfty/bmbl5/bmbl5toc.htm

20. Overview on human cancer cell lines20