PSIO 431 Test 9 and 10, 11 and 12
Immunological techniques for measuring antigens
Same idea as Elisa with a difference, measuring antigen not antibody. Well on an Elisa plate with an antibody on the bottom of the plate. Serum with antigen added, add a second antibody. darker the solution will be with a peroxidant - antigen captured by 2 antibodies - capture Eliza - antigen sandwiched
News article
Scientist did CRISPR genome editing and removed CCR5 from babies. Protecting them from HIV. But we don't know what happens when you mess with a genome like this. Other story: women didn't know she was HIV positive until giving birth. Many babies are infected during birth. Does not cross during placenta. No chance to treat mom prior to birth. What they did: started newborn on high dose of HAART. treated baby for 4 months, thought they cured child. Mother stopped brining child in for treatment and went off grid. Baby was showing antibody a year later, ended up not working.
Small Pox
Small lesions, discomfort, after blind 50% mortality 300-500 million death - associated in vessels 1967- 1 million cases every year 1980 - eradicated last natural case - 1977 reporter went to small pox lab and vial broke-1978 2 places in the world that still have small pox around: Russia and the US Both hold onto it as a bio weapon - don't want to get rid of it in case they use it so we can make a vaccine
Discuss: Are tumor cells antigenically different from the normal cells from which they arise?
yes - something has changed that has given us a target
How the immune system kills tumor cells
• 1. Cytotoxic T cells. These are probably the most important cells in tumor resistance, and they're every right-thinking immunologist's favorite cell. • In the past decade it's been found that ►CTL have at least two "checkpoint inhibitor" surface receptors which, if engaged by corresponding ligands on the cell they are interacting with, signals downregulation of their cytotoxic activity. - tumor cells are clever, they down regulate MHC1 and up regulate "off signals" where they can turn off CTLs 1st: stimulation then inhibitors - turn it on, then slowly turn it off CD28 in the lymph node CTLA-4 in lymph but slower PD1: periphery 3 • 2. Th1 cells. • Discuss: How can we make people with tumors get this system going better than it is? - give an anti Il-4 antibody against Th1 • 3. Natural Killer (NK) cells. provide a link between the innate and adaptive immune systems PRRs = DAMPs Receptors for MHC class I NK bind to MHC 1 = suppress response - looking for cells that down regulate MHC 1 Receptors for IgG = complement activation and components = rid of cancer cell CTL and NK: cops drive through neighboorhood, CTL is looking if the mail is right NK is looking for lack of MHC and making sure everyone has their porch light on - finds they can both kill directly
DNA vaccines
• A number of groups are investigating the possibility of immunizing not with the antigen, but with DNA that codes for it. - take DNA, inject it into you and now you make your own antigen • This DNA, isolated from the infectious organism and cloned into a suitable expression vector, would be injected into the skin or muscle. • There it would get into cells, and be transcribed and translated. • You'd become your own source of antigen, for life. • The advantage is speed: a vaccine like this could be available in a couple of weeks,
Rapid screens
• A throat swab is extracted, and the extract passed through a membrane to which Strep antigens stick because there's a dot of anti-Strep antibody coupled to it. • To detect this binding, the kit contains antibody against Strep to which liposomes (little fat droplets with a water interior) have been bound; the water contains a dye. • This preparation is also passed through the membrane, and it sticks if antigen has been trapped by the dot. • Something is added to pop the liposomes, and so the spot turns color if there was Strep antigen in the extract.
How it started
• AN OUTBREAK OF A RARE PNEUMONIA... • In early June of 1981 Gottlieb and co-workers recorded the cases of five young men, previously in good health, who were treated for Pneumocystis jirovecii pneumonia in Los Angeles hospitals
Discuss: You believe a person has a bacterial infection. How would you test this?
- see whether they have antibody against bacteria - do they have the antigen If we are looking for bacteria itself, we test for the bacteria = direct test If we think they have bacterial infection but we test for antibody = indirect test
HIV Entry into Host Cells
1. HIV (red) with Gp 120,41 blown up so were can see whats happening. 2. CD4+ cell = T helper cells are at risk of infection of HIV. 3. GP120 binds to CD4, when they bind a conformational change occurs 4. GP120 binds to another surface transmembrane protein called CCR5 5. GP41 binds to cell membrane and melts membrane allowing what was inside HIV RNA capsule to enter the cell
Think about and Discuss: From easiest to hardest, list tests you can think of to measure B cell function.
1. Look for B cells - nothing about function 2. Look for antibodies in the blood - know our B cells can make antibody 3. What classes of antibodies - breakdown is important 4. Look for specific antibody - antibodies present means you've been exposed 5. Lymph node biopsy (painful) - look at cortex
Discuss: What's the story with the gamma globulin peak in each of these traces? Which (if any) is normal?
1. No gamma peak = we are missing antibody = agammaglobulinemia = not normal = Bruton's disease 2. Large gamma peak = making a lot more antibody = polyclonal hypergammaglobulinemia = lots of B cells making antibody = Normal response to infection can also be leukemia 3. Gamma finger Monoclonal hypergammaglobulinemia = single type of B cell making antibody = antigens in general are polyclonal so this is not normal = form of cancer called multiple myeloma not the most sensitive test but cheap can only really be used for certain antibodies, not IgA
Passive Agglutination 3
Illness suspected and looking for pathogen. Beads with antibody, take serum for individual and look for agglutination Have to do a spinal tap to get fluid and add it, if bacteria Is there it sticks and falls out of solution. seeing if they have the antigen
Discuss: List all the steps that must take place to get a positive test.
1. Paint skin 2. Soak down deep to get eaten by dendritic cell (recognized and eaten) 3. Presented to proper T cell via cross presentation 4. Clones of DNFB recognizing cells are made 5. Memory cells are honed to the next spot and secrete proper cytokines 6. Macrophages call in, activate, and eat via M1. 7. Macrophages cause response Others; Stimulate T cells in mitogen and look at cytokines Thymic scan Biopsy of the lymph node, easies t is the rectal mucosa, easy to get to and limited pain and nerve endings
Discuss: If your pharmaceutical company was developing an immunotherapy for treatment of cancer, what would it look like?
1. make sure we are activating proper cells, don't want nonspecific cells CTLs and TH1 and NK 2. Want to expand numbers, personalized 3. Memory? Monthly infusions of antibody and when does it stop
Cause
AIDS is caused by a virus called HIV-1, for Human Immunodeficiency Virus. 2019-New strain (first in 19 years!!) - not causing much issue :) HIV is a nontransforming retrovirus, that is, an RNA virus that carries no oncogene, and reproduces itself by copying its RNA into DNA by means of its enzyme, reverse transcriptase. - injects RNA into our nucleus and creates its own reverse transcriptase - HIV jumps species; jumped from monkeys (SIV virus) via contact with monkeys that we didn't have before or people may have even eaten the monkey meat electronmicrograph of HIV**
Pathogenesis
After a single exposure, infected people develop high virus levels, and then antibody to HIV in 3 to 9 weeks. People who are antibody-positive (seropositive) develop AIDS symptoms at the rate of about 3% per year. From HIV infection to AIDs is about 9.5 years The mean incubation period is thought to be about 9.5 years How did they figure this out? - Need to know exact exposure dates, difficult. - Followed infected people through blood banks, knew exactly when individual received HIV. Reason that gay people can't donate blood STILL.
Testing infographic
After positive test: - continue by doing an immunoassay - western blot looking for HIV associated proteins - RNA tests; Northern blot - ELISA
Passive Agglutination 2
Agglutination at bottom of test tube titer of antibody in this examples = 1/64
Discuss: How would we get rid of a Viral infection?
Inside cells 1. Need Th1's and CTL's - virus gets in body but has not infected cells = antibodies can prevent viral infection - T cell immunity necessary for recovery
Passive Agglutination 1
Antigen attached to beads - serum added and antibody will bind and fall out of solution via complement - lattice falls out of solution - take serum and see agglutination, need to see how much we have Now look for titer of antigen 100% serum, dilute and see if you get agglutination, see where it stops
Flow Cytometry
Best way to measure 1. Test tube with cells (T cells) have added antibody against CD4.. put a color tag on it, add CD8 with different color 2. Small straw comes down and pulls cells up one at a time 3. As it goes up straw, laser shines on the cell and scatters, can get information from scatter on size of cell and granularity. - Bright yellow or red then its CD4 or CD8 4. Plots based on size and florescence - dots dependent on # of cells
Discuss: A baby has a 64 titer of IgG antibody against rubella at age 8 days; another has a titer of 64 at 8 months. In each case, how do you interpret it? (The minimum titer for protection is about 10...that is, the serum can be diluted 1:10 and still neutralize the virus)
Both have a titer of 64 but need 10 - baby are still protected and have a high enough titer 8 days: Mom made IgG against rubella, need to know if IgM is signaling = infected 8 month: Making own IgG, baby is infected
Discuss: Flow cytometry results on single cell suspensions prepared from biopies of 2 lymphoid organs, stained with 2 Abs with different fluorescent tags.What cells are 73% (A), what cells are 3% (A), what cells are 4% (B) What are A and B biopsies of?Are these normal or abnormal biopsies?
Brighter the color the more cells - all have CD4 and CD8 tag X-axis: CD4 Y-axis: CD8 What cells are 73% (A): Have both CD8 and CD4 = THE THYMUS! Only place we should see double positive what cells are 3% (A): neither CD8 or CD4 = probably epithelial cells or any other cell what cells are 4% (B): Neither CD4 or CD8 = THE LYMPH NODE = 4% CD8 positive CTLs (CD4 Helper) What are A and B biopsies of? A = Thymus B = lymph node Are these normal or abnormal biopsies? A = normal B = normal CD3 to know all T cells
Incidence
By 2004 AIDS was the 5th leading cause of death in the world, the 4th in developing countries. So far, 84.2 million have been infected (2018) and over 40 million have died. Currently- 38.4 million people with HIV/AIDS Approx 79% of people with HIV knew their HIV status in 2018
Equilibrium
Equilibrium. In most clinically relevant tumors, T cells infiltrate the tumor, but do not fully destroy it. Instead the tumor and T cells exist in an equilibrium.
Escape: the tumor cells fight back.
Escape: the tumor cells fight back. If a patient's tumor regresses (goes away), did the immune system have something to do with it? Melanoma patients, broke into two groups: - both treated w surgery regressors: tumor went away Progressors: tumor grew isolated T cells in the groups and exposed them to melanoma
ELISA-enzyme-linked immunosorbent assay
HIV detection Well turned sideways, in each well we attach or purchase antigen stuck to the bottom. Now we look to see if there is antibody against the antigen by taking some serum and adding it to the well. If antibodies present they will bind. Wash off free floating antibody and measure the stuck antibody. We need a way to measure it by putting a a second antibody that recognizes the second chain. Quantifies amount of fluorescence.
Think about and Discuss: How do you think our perspective of HIV/AIDS has changed in the last 40 years?
HIV is one of the few conditions where the virus causes a disease with a different name.. AIDS - Covid 19 is the disease caused by Sars-CoV2 Who gets it? Has changed, first started that homosexual males only got it. Now all around. How its transmitted. Transmission has been discovered, when its first started people didn't want to be in contact with a person with HIV How its treated? Don't yet have a cure but have medications that decrease symptoms to nothing and keep people alive, originally a death sentence. Can also pretreat.
Discuss: What are some issues with creating a vaccine for HIV?
HIV may be the closest thing to a perfect virus we have encountered. It is variable, it destroys the patient's defenses, it can hide, it allows time for spread before it kills. Fortunately, it is not highly infectious. It presents an incredibly difficult intellectual and moral challenge to scientists, physicians, and people.
Discuss: How do you think physicians and scientists were able to determine that AIDS was a new disease (rather than an old disease that was reappearing)?
Has to do with the pneumonia, because its rare, every time a person presented with it it was sent to the CDC. They were able to use history to look for this disease. Right at 1981 the numbers went way up.
Discuss: Why do you think CD19 or CD 20 is more specific than IgD or IgM for measuring B cell numbers?
IgM not a great way to measure because IgM can also be secreted into the serum = abnormally high # IgD is only a B cell receptor but it can also be secreted and free floating - more specific to use something that never comes off surface
Tissues Infected by HIV
Lungs: Low IgA Lymph: Low CD4 Brain: HIV associated dementia
Discuss: Cancer seems to be a disease of wealth (it's not in the top 10 killers in the world's poorest countries). Why do you think this is true?
More developed = top 10 killer. - poor countries might not live long enough to develop cancer - age is proportional - access to treatment - different exposures - different nutrition and activities
Measles
Must have high population immune for it to not be a risk people stopped vaccinating and problems began occur. Huge outbreak at Disney land in 2015 vaccine hesitancy
So This Bobcat Walks Into A Bar
No, really: COTTONWOOD, Ariz. (AP) The bobcat came in, patrons got bit and scratched, needed to be tested for rabies. Any wild animal can cary and its still very serious. Bats, skunks, coyotes 1. Go to the doctor and hopefully bring the animal - beneficial to know if they had rabies or not
Discuss: In what way are radiotherapy, chemotherapy, and surgery not specific compared to immunotherapy?
Not going to get all the cancer cells - big risk of other issues - chemotherapy destroys RBCs - immunotherapy: turning on immune system and too much activity can lead to autoimmunity
Evidence for cancer immune surveillance
People with immunodeficiencies, particularly of T cells, have a higher incidence of tumors - AIDs - organ transplant have 25-100 fold increase in tumors - nude mice lack T cells and rarely get tumors Activated T cells that recognize tumor- associated antigens can be identified. A small percentage of tumors, mainly melanomas, spontaneously regress, presumably due to an immunologic response. - cancer is a failure of immunity
Cause cont.
Picture of HIV Outer membrane has some proteins called gp120 and gp41 - RNA is inside capsule with its own reverse transcriptase ^ targets for intervention No vaccine because: This is the most antigenically variable pathogenic virus we have encountered. (In a single individual that have 1000s of variants of HIV, not effective) Reason: Reverse transcriptase is a highly error- prone enzyme, without proofreading capability. - makes mistakes and changes up frequently It makes a mistake about 1 in 100,000 bases, so everybody who is infected will have many variants in their body.
Experiment #3
T cells (Progressors or regressors) + serum from progressor patients Result: No killing Blocked T cells from killing tumor - cytokines - some tumor cells down regulate MHC Class 1 - no MHC then the CTL doesn't know what to do
Experiment #2
T cells from patients with growing tumors - progressors - T cells did kill melanoma - surprise!
Experiment #1
T cells from regressors - tumors went away - bottom of the dish has melanoma cells - add T cells - T cells were able to kill melanoma cells conclusion: great! expected
Discuss: What types of infections would you guess are seen in patients with AIDS?
Think about Helpers! Th1 - no Th1 = no help to M1's or CTL = increase in viral infections Th2 - no help to M2's, the clean up crew = decreased cleaning of wounds Tfh B - help B cells = antibody = no antibody = bacterial infections, parasitic infections Tfh 7 - See Th1 Treg - Could see increase cytokine = issues - everything but innate is affected
Immunodiagnosis
This refers to two things: the diagnosis of immunological diseases, and the use of immunologic techniques in the laboratory for the diagnosis of other conditions. Most of the methods used in a big reference or hospital laboratory (Banner) are described here. Lymphocytes: CBC = complete blood count - white cell total - platelets - RBC Next, differential: # lymphocytes and if cells # are normal - computer does it easily and quickly
Tumor antigens
Umbrella • TAA: Tumor associated antigen, also found on normal cells but likely a quantity thing. don't want to target • TRA: Tumor rejection antigen (is a TAA) leads to rejection of a tumor, recognized and destroyed. • TSA: Tumor specific antigen (TAA) only found on super specific tumors - chemical/ physical - burns - smoking • Oncofetal antigens: (OFA): found on fetal tissue, not normal on adults - carcinoembryonic - adult = colon cancer • Differentiation antigens - differentiation of different organs - over expressed antigens if cancerous - prostate specific antigen • Clonal antigens - specific clone - surface markers - B cell malignancy - look at Ig idiotype
...and a rare tumor
Very soon after this publication came another about 18 young homosexual males from California with a peculiar tumor, Kaposi's sarcoma This was disturbing: Kaposi's sarcoma was, until then, one of the rarest tumors known. About 25 cases had been reported annually in the USA, and almost all of them were in elderly men. As it became obvious that homosexuals were not the only group at risk, the disease was renamed AIDS, for Acquired Immune Deficiency Syndrome 3 patients with hemophilia needed blood transfusions and got AIDS
HIV Life Cycle and Sites for Therapeutic Intervention
Where we can intervene: Entrance inhibitors: try to block from entering cell Reverse transcriptase inhibitors: keep transcription from happening Integrase inhibitors: keep viral DNA from integrating into our DNA Protease Inhibitors: Blocking it from entering HIV core structure As of right now, most common treatment is HAART reverse transcriptase and protease inhibitors combined
HIV Life cycle
Within cell, RNA enters and is converted by reverse transcriptase into DNA, enters nucleus and integrates with our DNA so we have our DNA and virus DNA. Then we translate to RNA, make protein, assemble RNA, and then bud off of Thelper and make more HIV We make antibody against HIV but not helpful... A very significant behavior of the virus is this: when the virus is replicating, gp120 is made first and it becomes inserted into the cell's plasma membrane. CD4 infected and puts up GP120, which binds to CD4+ cell. Easily we form syncytium = group of two things acting as one. Allows HIV to move from one cell directly to the other in one phase. This apparently allows fusion of the infected cell to nearby CD4 cells, and a syncytium forms. In this way the virus could spread without an extracellular phase. This could easily explain why the antibody patients make seems to be useless.
Discuss: Smallpox eradication is a true success story. Why do you think we were able to get rid of this disease, but not others such as Polio?
Worldwide, the whole world got involved. Countries halted wars to vaccinate people Since then we haven't come together to rid a pathogen
Discuss: What are the disadvantages of DNA vaccines?
You have a cell and inject DNA, cell sticks a piece up on MHC Class 1 - CTL comes by and says "why is a liver cell in the thymus".. die. - risk of killing off our own cells - can see skin damage and muscle wasting
Discuss: What's the approximate concentration of IgA in the patient's serum?
between 320 and 160 somewhere around 200 mg/dL
Discuss: If this is true (equilibrium), what causes the cancer to "reactivate"?
chronic inflammation
Direct Immunofluorescence Indirect Immunofluorescence
direct: see if bacteria is there, throat swab. Buy antibody known against it with a fluorescent tag. If it lights up, yes. indirect: take a serum sample and purchase the antigen, add patients serum. then have to buy another antibody with fluorescent tag.
Discuss: How would we get rid of extracellular bacteria? Intracellular bacteria?
extracellular: majority of bacteria, try to make a vaccine and pump out antibody intracellular: much less common, microbacteria (TB, listeria) get into cells like macrophages and can stay in just fine. But if activated by Th1 it will kill anything inside it
Elimination
from one cell to one gram of cells (detectable tumor) is 31 generations of division. - cell divides every 24h then it takes around a month, but usually longer.. around 20 years. Cell is cancerous, new mutation and pink escaped, starts developing and keeps missing the killing of tumor by the time a new mutation occurs DAMP = damage associated molecular power MHC 1 = cancer antigen APC = macrophages come takes awhile to develop a cell so in order to survive it must change enough to not be recognized by our immune system
Discuss: What's the difference between Herd Immunity and Herd Effect?
herd immunity: the % of the population with immunity - must be high in order for it to work - what is the herd immunity we need, assuming infection makes you immune after herd effect: decrease in infection rate in the non immune portion of the herd
2 cases: cured
success story until recently, passed away. Berlin patient was HIV positive and had leukemia. Treatment; bone marrow transplant. In both cases their match did not have CCR5 (CCR5 deficient) 10% of people of Eastern European decent have this mutation. Bone marrow match CCR5 deficient and HIV can't affect because it has nowhere to bind next. They were cured of leukemia and of their HIV. Cant do it for everyone because of HLA, have to find a great match for it to work.
Passive antibody therapy
• Antibody to TAAs should be useful, and quite a few monoclonal antibodies (mAb) are already available. • They can be used as-is or they can be tagged with a poison such as ricin, or diphtheria toxin, or a radioisotope (such modified antibodies are called immunotoxins). put something deadly on antibody and when recognized it will die, can be very specific. treatment for prostate cancer
B cell numbers
• B cells in blood can be measured by counting cells with surface IgM or IgD. • Most labs use instead the markers CD19 or CD20 because they are more specific (Cluster designation) fluorescent tagging
Innocent bystander killing
• BCG is injected directly into the tumor. - • A ferocious delayed-type hypersensitivity reaction to BCG ensues, and the tumor cells are killed the way lung is killed in TB. solid cutaneous = melanoma bladder cancer
Ebola-Vaccine
• Ebola is a rare and deadly disease caused by infection with a strain of Ebola virus. The 2014 Ebola epidemic is the largest in history, affecting multiple countries in West Africa. The risk of an Ebola outbreak affecting multiple people in the U.S. is very low. • Prevention of Ebola viruses from infecting humans is the best way to protect individuals 2 vaccines in trials 2 more in development
Rabies
• For rabies, active immunization is possible with a vaccine grown on human diploid cells (HDCV), which has admirably few side effects. - can be vaccinated after the fact; starts slowly • If someone has been bitten by an animal known to be rabid, passive immunization with human rabies immune globulin is imperative, as well as active immunization (6 to 8 shots of HDCV). - also treated with immunoglobulin • Because the onset is slow, this is one of the few diseases where immunization may begin after exposure • Otherwise,the case fatality rate is 100%. - some exceptions - 6 survivors but all have residual neurological damage - 5 the vaccine didn't work wheel chair girl: 15 years old, found a bat and picked it up and bit her. Mom treated it, no doctor 1 month later symptoms: slurred speech, neurological disorders.. had rabies - put her in a coma to slow down brain activity - pumped her with antibiotics for 3 weeks - survived but has some neurologic issues
Hodgkin's lymphoma
• HODGKIN'S LYMPHOMA is a mystery, which centers around an abnormal, probably malignant, cell in lymph nodes (the Reed-Sternberg cell, often binucleate with prominent nucleoli) which is usually a B cell and may be carrying EBV genes. Epstein barre owl eye
Cancer
• It has been estimated that a human will undergo up to 10^16 mitoses in a lifetime. • Discuss: What cells do you think keep us from developing cancer constantly? - cells become cancerous, don't work like normal cells - cells that can recognize our own cells - CTLs (Killer Ts) - Th1 for help
Lymphoid malignancies
• LEUKEMIAS AND LYMPHOMAS. • When many abnormal cells are found in the blood, the condition is leukemia; if in the tissues, including lymph nodes and bone marrow, it is lymphoma.
Viral gene products
• Many tumors are known to be caused by tumor viruses; in humans about 20% of tumors are caused directly or indirectly by viruses. Virus and then develops a tumor HPV: human papilloma virus, big worry is cervical cancer Hepatitis = liver cancer Epstein Barr = B cell lymphomas
Prevention
• PREVENTION. Safe sex, safe addictions. • The virus is not hardy, and common disinfectants kill it readily. • PrEP: Pre-exposure prophylaxis • Only a vaccine will ever be able to put an end to this terrible worldwide epidemic.
T cell numbers
• T cells in blood get measures using CD3, CD4 and CD8
T cell function
• The best overall test: skin test with common antigens to which most people will have DTH. • A good set is: Candida, streptokinase/streptodornase (SK/SD,) trichophytin, mumps, tetanus, tuberculin. Read at 24-48 hours. challenge DTH = paint substance on skin, don't wash off. Use DNFB = 98% of population will become sensitized. Look for response in 10 day and T cells will make response
chronic lymphocytic leukemia (CLL)
• The most common B cell disease is chronic lymphocytic leukemia (CLL) • It is a malignancy of resting B cells, which are sIg+ but rarely secrete immunoglobulin. • The prognosis is good, because these cells (or their precursors) divide slowly and secrete no harmful products.
Diagnosis
• The most common test is for antibody to HIV.
Specific immunization
• There hasn't been much success in this area yet, but as more defined TAAs are being prepared, keep an eye out for breakthroughs. Taking out tumor cells, adding a gene that increases antigen on surface, reimplant - more or less epitopes want more
Antibody titers in diagnosis
• Titer, incidentally, is the reciprocal of the maximal dilution of a patient's serum that is still positive in some defined test. • So...if I can dilute your serum 1/64 and still find antibodies (that can agglutinate or precipitate) some antigen, my titer is 64. German measles, rubella is a good example of measuring titers -pregnant = can cross placenta - infected fetus has 75% risk of developing congenital issue - give pregnant late
single radial immunodiffusion
• To measure levels of individual immunoglobulin classes or subclasses we can use single radial immunodiffusion - immunoprecipitation but in a gel not a test tube. - in gel your interested in IgA, then Gel has anti-IgA - poke holes in the gel into one and put standards of IgA - serum in whole diffuses into the cell - reach equivalence = lattice around the well and its where equivalence is reached
B cell function
• To test the humoral arm of the immune system, we begin with serum protein electrophoresis: Make a gel, put some serum, move from negative to positive. Smaller the protein the farther it moves - scan protein and find out how wide and dark it is - focus on the gamma peak
Tests for specific antibody
• We often want to measure antibody to a particular antigen, as opposed to total levels of immunoglobulin, as evidence of current or prior infection or immunization with a particular pathogen. • How would you do it? If we know what the antigen is, we mix serum with antibody and see if = precipitation = not super sensitive aggluntation test = antibodies on beads
