2. GENERALIDADES
LEUKEMIA is a neoplastic disease that
occurs as a result of unregulated proliferation
and incomplete modulation of cellular precursors
of hematopoietic and lymphoid tissues.
In BONE MARROW tumor cells displace the
remaining normal series and later spreads to
other sides of the body:
Lymph-nodes
Liver
Spleen
3. GRENERALIDADES
It is a type of cancer that begins in the white blood
cells in the bone marrow.
Over time these cells tend to accumulate and many
people with the disease do not show symptoms for at
least a few years.
We have discovered two types:
-one fast progress.
- one slower progress.
4. GRENERALIDADES
B lymphocytes (cells that protect the body
against invading germs by maturing into
plasma cells that produce antibodies) are the
cells that are affected more frequently in CLL.
SYMPTOMS
-Weakness
-Sensation of fatigue
-Weight-Loss
-Fever
-Enlarged lymph-nodes
5. INTRODUCTION
MOLECULAR MARKER is a character or a gene may be
used to indicate the presence of another gene.
TN ANTIGEN is the first product GalNAc transferase.
GALNT11 is a variety of the 14 GALNTs this overexpressed
in patients with CLL, in B cells.
6. INTRODUCTION
THE ARTICLE IS BASED ON
RECENT DEVELOPMENTS.
SUCH US
THERE ARE TWO PROFILES AND A WIDE RANGE OF
DIFFERENT MANIFESTATIONS.
•(IGHV) MUTATED (M) ---> BETTER PROGNOSIS
•(IGHV) UNMUTATED (UM) --> WORSE PROGNOSIS
THIS ANALYSIS IS COSTLY AND DIFFICULT
8. INTRODUCTION
ONE OF THE MOST IMPORTANT
ALTERATIONS IN CANCER CELLS
THEREFORE THE EXPRESSION OF THE
ENZYMES OF THIS PROCESS IN CANCER
CELLS (GALNTS) WAS INVESTIGATED.
SPECIFICALLY GALNT11
EXPRESION OF RESULTING OF AN
EXPRESION OF RESULTING OF AN
INCOMPLETE O-GLYCOSYLATION
INCOMPLETE O-GLYCOSYLATION
TN ANTIGEN
TN ANTIGEN
9. GENERAL OBJETIVE
THE GOAL IS TO STUDY GALNT11
AS POSSIBLE NEW MOLECULAR
MARKER IN CHRONIC
LYMPHOCYTIC LEUKEMIA.
TO USE IT AS A DIAGNOSTIC TEST
FOR THIS SPECIFIC TYPE OF
LEUKEMIA AND BUILD MORE
APPROPRIATE AND SPECIFIC
11. MATERIALES Y MÉTODOS
SANGRE PERIFÉRICA
CONTROLES
SANOS
PACIENTES
CLL
CENTRIFUGARON
PURIFICARON
CELULAS T
CELULAS B
12. MATERIALES Y MÉTODOS
EXTRACCIÓN DE RNA
SE TRATÓ CON AMBION
(TRATAMIENTO LIBRE DE DNA)
MMLV
(TRANSCRIPTASA REVERSA)
VARIOS CICLOS DE PCR
CONVENCIONAL
RT-PCR
PARA IDENTIFICAR LA EXPRESIÓN
DE DIFERENTES TEJIDOS
LPL
GALNT
13. MATERIALES Y MÉTODOS
QUANTITATIVE REAL TIME PCR (q-PCR)
SYBR GREEN (SUSTITUTO DE
BROMURO DE ETIDIO) TINCIÓN
GALNT11
FLUORESCENCIA EN RELACIÓN A LA
CANTIDAD DE ÁCIDOS NUCLEICOS
PRIMERS
AMBION Y RETROTRASNCRIPCIÓN
AMPLIFICACIÓN
GAPDH (GEN DE
REFERENCIA)
CD3 (MARCA DE LINFOCITOS T)
CONTROL DE CONTAMINACION( 6 DONANTES SANOS)PCR POR DUPLICADO(TODO) FUSION DE CURVAS
14. MATERIALES Y MÉTODOS
PROTEINAS DE UNION ESPECIFICA Y
REVERSIBLE A CARBOHIDRATOS
ANTICUERPOS Y LECTINAS
SE FORMO
ANTIGALNACANTIBODY DE
CONEJO
DETECTA PROTEÍNA
RECOMBINADA DE GALNAC-T11
MAB B72.3 (AC)
MAB 83D4 (AC) IGM
VVB4 LECTIN (SIGMA)
DETECTAN PRESENCIA
DE TN ANTIGENO
15. MATERIALES Y MÉTODOS
EXTRACCION DE PROTEINAS,SDS PAGE, WESTERN BLOT
GAPDH AC FUE USADA COMO CONTROL
EN ESTA EF SON DISUELTAS EN
SDS(DETERGENTE DUODENIL SULFATO ) CARGA
NEGATIVA
SEPARACION DE PROTEINAS SEGUN
TAMAÑO POR EF EN PAGE
CUANTIFICACION DE PROTEINAS POR BCA-ENSAYO
FILTRO
CUANTIFICACION RELATIVA POR
GENTOOLS
QUIMIOLUMINISCENCIA
INCUBACION CON AC
16. MATERIALES Y MÉTODOS
CITOMETRÍA DE FLUJO
INMUNOFLUORESCENCIA
IDENTIFICA INCLUSO PEQUEÑAS
POBLACIONES CELULARES
UNION AC CON MOLECULA
FLUORESCENTE ( ABSORBE LUZ UV)
TRANSCRIPTASA INVERSA
CON ALTA ESPECIFICIDAD
A PARTIR DEL CD Y MIDE VARIOS
PARAMETROS
DETECCIÓN DE HORMONAS Y
PROTEÍNAS
PBS( BUFFER FOSFATO SALINO)
REACCION ANTIGENO-AC
21. RESULTS
POSITIVO
- CLL
- CÉLULAS T
- JURKAT
NEGATIVO
- CÉLULAS B
- DAUDI
CONTROL NEGATIVO
Intensidad
Intensidad
depende de la
depende de la
concentración
concentración
de la PCR
de la PCR
35. BIBLIOGRAPHY
- MARTINEZ SÁNCHEZ, Lina María. (2012) . Class book , Molecular Biology
, 7 ed. Colombia, Medellín: UPB Medical Faculty, Editorial Universidad
Pontificia Bolivariana pp.7-8.
- M.G. Libisch, M. Casás, ML. Chiribao, P. Moreno, A. Cayota. (2013
September 17). GALNT11 as a new molecular marker in chronic
lymphocytic leukemia. Elsevier B.V. Retieved from:
http://www.sciencedirect.com/science/article/pii/S0378111913012572