T-Cell Testing Made Easy
Nutritional immunologist Tim Kramer examines a collection of individual T-cell
proliferation cultures harvested onto the filter mat of a 96-well cell
harvester. The samples can be used to determine how much T-cell growth has
The need for a quick and easy way to test people's immune competence first
hit Tim Kramer in 1976 while working at the Anemia and Malnutrition Research
Center in Chiang Mai in northwest Thailand.
Kramer was returning a previously malnourished child to her home some 4
hours' drive from the center, over very hilly terrain. As he passed through the
small villages along the way, he thought, "I'd love to have some data on
how malnutrition affects cellular immunity in such a location."
In layman's terms, that's the ability of T-cells to rapidly divide when they
bump into a chemical stimulus, or antigen. Among the immune cells, Kramer
explains, T-cells are the most sensitive indicators of a person's nutritional
status, because their activity is altered by even marginal nutritional
But it would have been very difficult technically to do such measurements in
a makeshift laboratory in the countryside, says Kramer. The traditional method
requires at least 10 milliliters of blood. Many people in Thailand and other
Asian cultures are opposed to giving blood. Assuming one could get enough, the
T-cells then would have to be separated by highly trained technicians using
expensive equipment before they could be cultured and subjected to a foreign
substance to prompt them to divide.
As a solution, Kramer thought, "Why couldn't we use a microculture of
whole blood instead of separating the lymphocytes [T-cells and related
cells]." Such a technique would require only a few drops of blood and
significantly reduce the technical requirements . . . and thus the cost.
But it took 11 years and Agricultural Research Service (ARS) backing before
Kramer had the time and resources to begin developing the whole-blood culture
technique. "And it's so simple, it's ridiculous," he says. Kramer is
a nutritional immunologist at ARS' Beltsville (Maryland) Human Nutrition
Refined, Over Time
For the last decade, Kramer has tweaked the technique into a foolproof
measure of cellular immune competence for nutritional studiesparticularly
for use in field studies of large populations. During that period, a handful of
other immunologists began developing the technique for other types of research
and clinical use.
"But it's still not widely accepted among immunologists," says
He hopes the situation will change after publication of his method by the
National Academy of Sciences. Because of its simplicity and reduced cost, it
could be used routinely to screen infants and children, the elderly, and others
whose immune competence may be suspect.
"The technique is ideal for pediatric use," says Kramer, since it
requires only 0.4 milliliter of blood or lessjust a few drops.
"That's one-twenty-fifth the amount needed for the traditional
And it costs 35 to 40 percent less because it doesn't require a microscope
or centrifuge and the technical know-how to use them. Instead, he says, a less
trained technician can easily handle three times more samples than with the
Immune cells, especially T-cells, are the body's main defense against viral
infections and parasites. They also play a major role in countering bacterial
and cancer cells before they get a toehold. By multiplying faster than the
invaders, immune cells keep them suppressed, says Kramer. "It's a numbers
What's more, T-cells coordinate the whole defense. That includes stimulating
the production of antibodies (immunoglobulins) by constantly
"talking" to one another and other cells, immune types and otherwise.
They talk through cytokinessmall chemicals with names like interleukin-2,
interferon-gamma, and tumor necrosis factor. And they listen through
receptorscomplementary chemicals for these cytokines on the surfaces of
cells. The body's immune response is a very complex scenario, says Kramer.
Not Only QuickerBetter
That complexity is why immunologists think they have to separate immune
cells from the blooda veritable chemical soupto get an accurate
measurement of immune competence.
Kramer says it took about 2 years of testing to convince himself that
whole-blood cultures could produce accurate results. Now he believes they give
a more authentic picture of what's happening inside the body, because the cells
are growing in a familiar milieu rather than among the foreign proteins in
Kramer's whole-blood culture technique passed its first shakedown in 1987,
when he was back in Thailand as an ARS researcher testing the effects of zinc
deficiency on the immune competence of children. Two years later, it withstood
the rigors of a study at the Beltsville center.
But it really proved its value in 1991 in Linxian, China, where Kramer was
cooperating on a large study pitting various antioxidant combinations against
cancer of the esophagusa major disease in that part of China. In just 3
weeks, he single-handedly tested some 15,000 whole-blood microcultures for
T-cell response. Side-by-side tests showed that the results were just as
accurate as those gotten traditionally.
Since then, he has used the technique exclusively in several nutrition
studies. It has shown a significant drop in the immune response of U.S. Army
Ranger trainees during periods of extreme food restriction coupled with
physical stress. [See also "Calorie Slashing and Overexertion Can Stress
the Immune System," Agricultural Research, January 1995, p.22.]
In a study of infant patients in a Washington, D.C., hospital, it
distinguished between those with active AIDS, those infected with the HIV
virus, and those with no infectionbased on the level of T-cell
"It provides a good in vitro mirror of what goes on physiologically in
people," says Susanna Cunningham-Rundles, associate professor of
immunology at Cornell University Medical Center in New York City. "What
Dr. Kramer has done is to optimize our ability to look at the interaction
between nutrients and the immune system in reality.
"Whole-blood cultures tell us whether or not the immune system is
altered," says Cunningham-Rundles, "but they don't tell us
howwhether it's due to changes in the number or types of immune cells.
That's where the traditional technique shines. But with technology being
developed," she says, "it should be possible to merge whole-blood
culture with the more analytical technique for routine clinical use and
And Kramer hasn't finished improving his technique. He says the chemicals
T-cells use to communicate among themselves also reflect the cells' activity
and thus indicate a person's immune competence. So he is perfecting the recipe
for getting accurate measurements of these cytokines and receptors. He's close
to finishing this work on two cytokines and is about halfway to optimizing the
technique for a third cytokine and a receptor. By Judy McBride
Tim R. Kramer is at the USDA-ARS
Human Nutrition Research Center, Bldg. 307, 10300 Baltimore Ave.,
Beltsville, MD 20705-2350.