Cast: Unni
Mukundan, Nithya Menon, Sweta Menon, Suraj Venjaramoodu, Siddique,
Baburaj, Manianpilla Raju, K. P. A. C Lalitha, Devi Chandana
Producer:
Director: T. K. Rajiv Kumar
Music Director: സരത്
തത്സമയം ഒരു പെണ്‍കുട്ടി ,കുറച്ചു പ്രശ്നജല്‍ ഒയിച്ചാല്‍ കണ്ടു എരികവുന ഒരു പടം ആണ് തത്സമയം .ടി.k .രാജീവ്‌ കുമാര്‍ ഒരു നല്ല സംവിധായകന്‍ യീന്ന നിലയില്‍ അദ്ധീഹം വിജയിച്ചു .എനാല്‍ രാജീവെന്ട കഴിവ് ആനിസരിച്ചു സിനിമ ആയില എന്തു പറയാത്ത വയ്യ .സിനിമ പാട്ടുകള്‍ നിലവാരം പുലര്‍ത്തി ചാനെല്‍ സംസ്കാരത്തിന ചോടിയം ചിയുക ആണ് സിനിമ 100 % entertainer ആണ് . ഇനി സിനിമയ പട്ടി
സിനിമഉട തല വാചകം കനികുംപോള്‍ ഉള്ള ചായ കട രംഗം salt ന pepper എന സിനിമ ഉട indrodution കിട്ടിയ പ്രക്ഷക പ്രശംസ അനുകരിക്കാന്‍ നോകിയ താനോ ഏന് തോണി പോകുന്നു എനാലും അതെ ആരാധകന ഇഷ്ട പടുതുനു .
പെനിട് സിനിമ കൊട് പോകുന്ന ഓരോ രംഗവും അകംശയും രസവും ഉണര്‍ത്തുന്നു .
സമുഹത്തില എല്ലാ അനിതികും എതിര സിനിമ camera zoom ചയിഉനു .എനാല്‍ vilapill shala മാലിന്യ പ്രശ്നം പകുട്ടിക് നിര്‍ത്തി കഥയ പുതിയ വായിതിരുവിളക് കൊടുവ്നഹു സിനിമയ രണ്ടാം പകുട്ടി ആരാധകന പിടിച്ചുഎരിതുനു .
എനാല്‍ രണ്ടാം പകുതി നനകുനത്തില്‍ സിനിമ യുട അണിയറ പ്രവര്തകര്‍ക് കയിഞ്ഞില്ല .തിരകഥ രണ്ടാം പകുതി ദുര്‍ബലം ആയി .
അഭിനയത പാതി രണ്ടു വാക്
നായികാ നിത്യ മേനോന്‍ നനായി അഭിനയിച്ചു പക്ഷ climax എലേല്‍ അഭിനയം മതിയക്ട പോയി
നായികാ തുല്യ പ്രദയം ഉള്ള റോള്‍ ചിത സ്വത മേനോന്നും നന്നായി അഭിനയിച്ചു
. നായകന പട്ടി പരുക അനന്കില്‍ പോര അഭിനയന്‍ .ചാനെല്‍ c .ഇ.ഓ ആയി തിളങ്ജനും ക്ലിമക്ഷെല് ഒരു നല്ല വില്ലന്‍ ആകാനും siddiqueഉ കയിഞ്ഞു ചാനെല്‍ producer ആയി തിളങ്ജന്‍ ബാബുരാജഏന് കയിഞ്ഞു .
പെനിട് എട് തു പറയണ്ടത് സുരജേന പട്ടി ആണ് വലിപേ ആയി പോയി .ത്യലോഗ് റിപീറ്റ് ആയി വരുനത്‌ അല്ലവസരം ഉണ്ടാകി .കോമഡി സ്റ്റാര്‍ എല actors അവരുട റോള്‍ നനകി .അച്ഛന്‍ ആയി അഭിനയിച്ച മണിയന്‍ പിള്ള രാജു നനായി ആ റോള്‍ ചിത് ടിനി ടോം തുടഞ്ഞിയ എല്ലാ തരജലും കല്കി .
സിനിമ മൊത്തത്തില്‍ കണ്ടിരികം.
ഗുഡ് not bad ,entertainer
ഇതു എന്താ സിനിമ വിലയിരുത്തല്‍ ആണ് ഇതു നെഞ്ഞല്ക് എങ്ങനാ ആണ് തോന്നുക ഏന് എനികരിയില ഇതു ആരും വിമര്‍ശിക്കാന്‍ അല്ല എന്താ അഭിപ്രായം പറഞ്ഴു എന ഉള്ളു .
Determination
of the blood group of the subject by ABO system and rhesus system.(Identify the unknown "blood types" of the synthetic
samples).
PRINCIPLE:
Binding of antigens to antibodies is
mediated by non-covalent bonds. The kinds of bonds that hold the antigen and
antibody together in this complex are the same ones that are generally involved
in producing the tertiary and quaternary structures of proteins, namely, ionic
bonds, hydrogen bonds, hydrophobic interactions, and van der Waals forces.
Interactions between antigen and
antibody involve non-covalent binding of an antigenic determinant (epitope) to
the variable region (complementarity determining region, CDR) of both the heavy
and light immunoglobulin chains. These interactions are analogous to those
observed in enzyme-substrate interactions and they can be defined similarly. To
describe the strength of the antigen-antibody interaction, one can define the
affinity constant (K) as shown:
Affinity K= [Ab–Lg]
‾‾‾‾‾‾‾‾‾
[Ab][Lg]
= 104 to 1012
L/mol
Greater the K, the stronger the
affinity between antigen and antibody. These interactions are the result of
complementarity in shapes, hydrophobic interactions, hydrogen bonds and Van der
Waals forces.
Antibodies are also used to help
our bodies find and destroy "foreign" cells such as tumors. Because
antibodies bind tightly to only one type of structure on the surface of cells
(antigen), they can also be useful for identifying different types of blood
cells.
Our blood type is determined based
on the presence or absence of two proteins on the surface of our red blood
cells (Type A and Type B). There are four possible combinations of blood types
namely: Type A, Type B, Type AB, and Type O (contains neither A nor B
proteins). This is referred to as the ABO blood type. In addition, red blood
cells have a Rhesus factor or Rh, which is either present or absent. If the Rh
factor is present, the cells are referred to as Rh positive.
Blood group depends upon types of
antigen present or absent in the blood.There are four types of blood groups
.Type of blood group depends on presents or absent of rhesus factor in blood
cells When antibodies are mixed with their corresponding antigens on the
surface of large, easily sedimented particles such as animal cells, erythrocytes,
or bacteria, the antibodies cross-link the particles, forming visible
clumps. This reaction is termed agglutination.
Blood types are determined by using antibody
reagents that specifically react with the A, B, and Rh proteins on the surface
of red blood cells.
MATERIALS REQUIRED:
Slide, lancet, toothpick, cotton.
Reagents : Anti-serum A, Anti-serum B, and
Anti-serum D, alcohol.
PROTOCOL :
1. Take a
slide and label A, B and D. 2. Spray
the left “ring “ finger with ethanol or
wipe it with an alcohol wiper and let it air dry. 3. Take a
sterile lancet and puncture the
fingertip. 4. Press
the finger from top to bottom and put
the blood drops on the slide. 5. Add a
drop of anti-serum A, B and D to the blood drops. 6. Mix
the antisera in with the blood using a separate toothpick.Place toothpicks in the biohazard waste. 7. After
several minutes, observe agglutination and determine the blood type.
To learn the technique of ouchterlong
double diffusion.
PRINCIPLE
Interaction
between antigen (Ag) and antibody (Ab) at themolecular
level forms the basis for several techniques that areuseful in modern day scientific studies and in routine
clinicaldiagnosis. These techniques are
either based on the use oflabeled reagents,
a tracer or immunoprecipitation. Ouchterlonydouble
diffusion (ODD) or double immunodiffusion techniqueis one of the simplest techniques extensively used to
checkantisera for the presence of
antibodies for a particular Agand to
determine its titre.
In ODD assays,
solutions of Ag and Ab are placed in adjacent wells cut in agarose gel and are
allowed to diffuse radially. The Ag and Ab concentrations are relatively higher
near their respective wells. As they diffuse farther from the wells, their
concentration decreases. An antigen will react with its specific antibody to form
an Ag-Ab complex. At one point their concentrations become equivalent and the
Ag-Ab complex precipitates to form a precipitin line.
KIT DISCRIPTION
In this
kit, an antigen and a test antiserum are supplied. Students will perform the
ODD assay to determine the titre of the antiserum, by using serially diluted
antiserum against the antigen and observing the formation of precipitin line.
The highest dilution at which the precipitin line is formed is considered as
the titre value of the antiserum.
KT09S :The kit is designed to carry out 15 ODD experiments.
Duration of experiment:Experiment is
carried out over a span of 2 days, approximate time taken on each day is
indicated below:
Day 1: 1 hour (Preparation of gel & loading of antigen and
antiserum)
Day 2: 20 minutes (Observation and Interpretation)
MATERIALS PROVIDED
The list
below provides information about the materials supplied in the kit. The
products should be stored as suggested. Use the kit within 6 months of arrival.
MATERIALS
QUANTITY
STORE
KT09S (15 EXPTS)
Agarose 2 g
2 gm
4°C
10X Assay buffer
20 ml
4°C
Antigen
0.2 ml
4°C
Test antiserum
0.5 ml
4°C
Glass plate
5 Nos
4°C
Gel punch with syringe
I Nos
4°C
Template
2 Nos
4°C
MATERIALS REQUIRED
Glassware :Conical flask,
Measuring cylinder, Test tubes.
Reagent :Distilled water.
Other Requirements:Micropipette, Tips, Moist chamber (box with wet cotton).
Note:
ØRead the entire procedure before
starting the experiment.
ØDilute the required amount of 10X
assay buffer to 1X with distilled water.
ØReconstitute the antigen vial with
0.2 ml of 1X assay buffer. Mix well, store at 4°C and use within 3 months.
ØReconstitute the antiserum vial with
0.5 ml of 1X assay buffer. Mix well, store at 4°C and us within 3 months.
ØWipe the glass plates with cotton,
make it grease free for even spreading of agarose.
ØCut the wells neatly without rugged
margins.
ØEnsure that the moist chamber has
enough wet cotton to keep the atmosphere humid.
PROTOCOL
ØBoil to dissolve 100 mg of agarose in 10 ml of 1X assay buffer.
Cool to 55°C.
ØPour 5 ml of the gel solution onto a clean glass plate placed on
a horizontal surface. Allow the gel to
set, it takes approximately 20 - 30
minutes.
ØPlace the gel plate on the template provided. Punch wells in the
gel with the help of a gel
punch corresponding
to the markings on the template. Use gentle suction to avoid forming rugged wells.
ØSerially dilute the test antiserum up to 1:32 dilution as
follows:
oTake 5 μl of 1X assay buffer in each of the five vials.
oAdd 5 μl of test antiserum into the first vial and mix well. The
dilution of antiserum in this vial is 1:2.
oTransfer 5 μl of 1:2 diluted antiserum from the first vial into
the second vial. The dilution in this vial is 1:4.
oRepeat the
dilutions up to fifth vial
ØAdd 5 μl of the antigen to the center well and 5 μl each of neat
(undiluted), 1:2, 1:4, 1:8, 1:16,
1:32 dilutions of antiserum into the surrounding wells.
ØPlace the plate in a moist chamber and incubate at room
temperature, overnight.
ØAfter incubation, observe for opaque precipitin line between the
antigen and antisera wells.
ØNote down the highest dilution at which the precipitin line is
formed. This is the titre value of the antiserum.
OBSERVATION
We can see the presence of precipitin line in 1:16 dilution. So the
titre value is 1:16.
പുതിയ പടം വീണ്ടും ഒരു മഹ്ഴാ
direction : clinton .m . yuvamela
story: ram.k pillai
screen play :nowshik api
producer: rakhi ram productions
associate : devan p hari.
editing: aji mappilai
വീണ്ടും ഒരു മഴ
1)What is the principle behind Latex
Agglutination?
When a sample containing the specific antigen (or antibody) is mixed with an
antibody (or antigen) which is coated on the surface of latex particles
(sensitized latex), agglutination is observed. he interaction between a
particulate antigen and an antibody results in visible clumping called
agglutination. Antibodies that produce such reactions are known as agglutinins.
Agglutination reactions are similar in principle to precipitation reactions;
they depend on the cross linking of polyvalent antigens. When the antigen is an
erythrocyte it is called hemagglutination. All antibodies can theoretically
agglutinate particulate antigens but IgM, due to its high valence, is
particularly good agglutinin and one sometimes infers that an antibody may be
of the IgM class if it is a good agglutinating antibody. Occasionally, it is
observed that when the concentration of antibody is high (i.e. lower
dilutions), there is no agglutination and then, as the sample is diluted,
agglutination occurs. The lack of agglutination at high concentrations of
antibodies which is called the prozone effect is due to antibody excess
resulting in very small complexes that do not clump to form visible
agglutination.
2)What is agglutination inhibition test?
If the antibody is incubated with antigen prior to mixing
with latex, agglutination is inhibited; this is because free antibodies are not
available for agglutination.In agglutination inhibition , the absence of
agglutination is diagnostic of antigen, provides a high sensitive assay for
small quantities of antigen. for eg : Home pregnancy kits includes latex
particle coated with human chorionic gonadotropin ( HCG) and antibody to HCG.
HCG is a glycoprotein hormone secreted by developing placenta shortly after
fertilization. The addition of urine from a pregnant women, which contains HCG
, inhibits agglutination of latex particles when the anti-HCG antibody is
added; thus the absence of agglutination indicates pregnancy.
3)Why is glycine-saline buffer used to
reconstitute the antigen and antiserum?
It is an
amino acid, one of the building blocks of proteins. Also can play a role of
osmoprotectant, helping organisms to withstand osmotic stress.pKa value of glycine is 9.78, hence
the effective pH range for glycine is 8.8 and 10.6.So the basisiyy will suit
for reconstituting the antigen and antiserum.
4)Why
are latex beads used for coating the antigens? Can we use anything else? Why?
Which would be a better option?
1.List the conditions under which you
would get a positive result in the reaction and the factors responsible for it.
2.Suppose you do the reaction and get a
negative result where you think you should observe a positive reaction. List at
least 5 factors which you would consider for trouble-shooting the experiment.
3.Suppose you do the reaction and get a
positive result where you think you should observe a negative reaction because
you know that the antiserum is not against that antigen. List at least 3 valid
scientific reasons for such an observation.
4.How can you test and justify if the
antigen has been coated on the latex beads? (Other than testing it with the
positive reaction by addition of antiserum)
5.What is the role of blocking buffer
in the experiment?
6.Can you think of at least 5
applications for latex agglutination in diagnostics, labs, industry or
research?
Thalsamayam Oru Penkutty
