tetragenus, pneumococcus, bacillus influenza, or Friedlander's bacillus. Since the first group of organisms will also grow well on blood-agar, it is obvious that whenever we are in doubt we always use this medium.

When glucose agar is used, the bacteria are isolated by making several dilutions and plating. When blood agar is used, the plates must be made at the time the blood is drawn, and subsequently inoculated by streaking the infected material over the surface by means of a bent glass rod or platinum wire. By both these methods isolated colonies are obtained, and pure cultures can be made by "fishing" out the different ones. and making subcultures on the differential media necessary.

Pure cultures are essential because:

1. The organisms must be identified in order to determine if they are pathogenic and also to know what temperature must be used to kill them.

2. Different kinds of bacteria have different rates of growth on the same medium, and some organisms grow well together, while others do not, so in mixed cultures we do not obtain the organisms in the proper proportions.

3. Almost all cultures from surface lesions are contaminated with saprophytic bacteria, which usually outgrow the pathogenic ones, and are, of course, useless for making a vaccine.

After pure cultures of the pathogenic organism or organisms are secured it is easy to prepare the vaccine. Only two precautions must be taken: 1. The pure cultures should not be contaminated during the process, and 2, the proper dosage must be obtained.

Method. Several agar or blood-agar slants are inoculated with pure cultures of the organism from which the vaccine is to be made. The bacteria are spread over the entire surface and allowed to grow in the incubator. Twenty-four hours is usually sufficient. Salt solution (0.9 per cent NaCl and 0.2 per cent trikresol) is added to each tube, and the bacteria are washed down by rubbing the colonies free from the surface with a platinum wire. The heavy emulsion is then poured or pipeted into sterile bottle containing glass pearls and thoroughly shaken, either by hand or, better, in a shaking machine. In this way the clumps are broken up and a uniform emulsion obtained. The mixture is then poured into sterile centrifuge tubes and whirled for a

few minutes to remove any particles of culturemedium which may be present. The supernatant emulsion is pipeted into a sterile tube, and is now ready to be standardized.

Standardization of a vaccine means the determination of the number of bacteria in 1 cc. of the mixture. There are several methods for doing this. Wright's is usually the most convenient. When using his technic, we compare the number of bacteria in the suspension with the number of red blood-corpuscles in human blood. We select some one whom we know to have about 5,000,000 cells in 1 cmm. A capillary pipet with a mark about one inch from the tip and several clean slides are the only apparatus necessary. One or two volumes (one volume equals amount of fluid contained in the capillary tube between the mark and the tip), of potassium citrate are drawn up first, then exactly one volume of human blood, and, finally, one volume of the bacterial emulsion. The fluids are immediately expelled on a glass slide, then drawn up and expelled again. This is repeated several times to secure a uniform mixture. Small drops are placed on each of three or four slides, and ordinary blood-smears made. These are stained with methylene-blue or Wright's blood-stain. When examined, we see that the slide has numbers of bacteria distributed around the blood-cells.

A total count is then made of 10 oil-immersion fields of both the red blood-cells and the bacteria. From this count we can determine the number of bacteria per cubic centimeter of the suspension. For instance, if there are 300 bacteria and 500 red blood-cells in the fields counted, we say 300: 50::x (the number of bacteria per cubic centimeter) :5,000,000. Therefore, x equals 3,000,000, the number of bacteria per cubic centimeter.

Since we have determined the concentration of the mixture, it is easy to dilute it so that we have 100,000,000, 2,000,000, or 500,000,000, in each cubic centimeter. For dilution we use the salt solution which contains 0.2 per cent trikresol, so that we have the same concentration of the preservative in all dilutions of the finished product. The vaccine is now put up in 1 cc. ampules, several for each dilution, and sealed.

These ampules are now sterilized by heating in a water-bath, usually at 58° to 60° C for fortyfive minutes.

These are tested for sterility by making cul

tures on appropriate media and by animal inoculation. If negative results are obtained, the vaccines are ready for use. They are labeled and kept in the ice box until injected.

Vaccine Versus Sera. In the treatment of infections it is necessary to decide whether the therepeutic sera or the bacterial vaccines are indicated. Vaccines, as stated above, are bacterial suspensions which, when injected, stimulate the organism to the formation of antibodies. Sera are fluids containing antibodies already formed, and are injected into the circulation to supply antibacterial elements without stimulating the bodycells to the production of these substances. Hence in the use of sera the antibodies formed in the body-cells of the horse are supplied to the patient, and a condition of passive immunity is established, lasting a few days or a few months.

There has been so much written on biologic therapy that it seems almost unnecessary to discuss the indications and contraindications for vaccines and sera. There is, however, some doubt in the minds of the average practitioners, and even some difference of opinion among the clinicians who use vaccines, just when the vaccines or sera are indicated.

Theoretically, sera should be given only in the toxemias, such as diphtheria or tetanus, but clinical experience has taught us to use sera in the acute general infections where the condition of the patient is such that the blood should be supplied with antibodies already formed, as in serum, rather than to tax the system by injecting bacterial vaccines, thus giving the organism the additional burden of making its own antibodies.

We believe that when the indication arises for the injection of vaccines, there is still one more question to be settled, namely, whether to inject the autogenous, the stock, or the mixed vaccines. We are convinced that the auto-vaccines are the agents par excellence of biologic therapy, and have made it a law that whenever it is possible to isolate a specific organism or specific organisms, the autogenous vaccines should always be made and given in a scientific, systematic, and fearless manner. Why?

1. Their use is most logical and scientific, because they produce the specific antibodies most suitable and most effective for the individual

case.

2. Clinically, they have given us the best results.

3. They rarely, if ever, produce harmful or annoying after-effects.

4. Their preparation is simple, requires very little time, and is by no means expensive.

The most serious objection which can possibly be raised is that the time required in preparing the vaccines may greatly hazard the ultimate results. In rapidly progressing infections due to the more common organisms, it has been our custom to give a suitable stock vaccine as the primary dose, while the auto-vaccines are being prepared. Stock vaccines are indicated only in those cases where the suitable auto-vacoines cannot readily be obtained; for example, in the case of acne, where it requires from six to eight days for the full development of the acne bacillus culture. We have used mixed vaccines, both auto and stock, where mixed infection has been clearly demonstrated. Their use does not necessarily signify the physician's inability to make an accurate bacteriologic diagnosis, nor need there be any hesitation on the part of the doctor because of the fact that they have been designated as unscientific. The fact is that there is a clear, positive, scientific indication for mixed vaccines in all mixed infections. We do not wish to infer that all bacteria found in a given infection must be injected with one puncture of the needle. Indeed, when treating acne, for instance, we have found that the best results are obtained by injecting the staphylococcus into one arm and the acne bacillus into the other at the same visit. should not hope to cure a case of tuberculosis with a mixed infection by injecting tuberculin alone, nor can one expect to eradicate a cystitis due to the colon bacillus and the staphylococcus by injecting the colon bacillus or the staphylococcus alone. Again, we have found that, in a great number of cases, especially the old chronic or sub-acute arthritides, neither the primary focus nor the causative organism could be demonstrated. In these one must obviously resort to a mixed vaccine. Having had a large number of just such cases led us to the use of a mixed vaccine containing as its predominating organism the most likely cause of the infection. For example: In those cases where the primary focus, as nearly as could be determined from the clinical history, lay in the respiratory tract, we have had our

One

bacteriologist prepare a vaccine containing pneumococci, staphylococci, and streptococci.

In the case of gastro-intestinal origin, we have used the bacillus coli, staphylococci, and streptococci.

Where the primary infection was was in the aural cavity or the various skin lesions, in fact, any lesion bordering on a low-grade pyogenic infection, we have had excellent results from the injection of a home-made pyocyaneus and staphylococcus vaccine. This may not seem particularly scientific, it may even seem empiric,-but it has given us results where all other types failed, and is justified on account of the clinical results, waiting a more exact and scientific procedure.

Clinical Considerations. As you all know, sera, up to the present time, are said to be indicated only in the toxemias, and that vaccines are indicated in the bacteriemias.

In the last four years it has been our custom to give the anti-streptococci serum in all those cases of acute general streptococci infections, acute streptococci peritonitis, and in the extremely toxic cases of streptococcus infections of the extremi

ties.

In a series of over 200 cases of this type each patient received from 10 to 80 cc. of the streptolytic serum, and with a fall of the temperature and a subsidence of symptoms in a majority of the cases. You will understand, of course, that the sera are discontinued as soon as the auto-vaccines are prepared.

Our experience with the sera in scarlet fever has been limited to a few cases, but even in these we noticed beneficial results and never any untoward after-effects. In a surgical clinic we rarely meet with diphtheria, but in the few cases we have treated the results have been most gratifying. We have, however, noted in a few cases the symptoms of anaphylaxis which were so fully discussed before this society two months ago.

The antitetanic serum is given in all cases of injury or gunshot wounds in which there is an open lacerated wound inflicted by machinery or implements which have been in contact with the dust or dirt. This is our routine practice, and has been used in a series of over 150 cases, and so proficient has been the prophylactic action of the serum that Dr. Murphy has not seen a case of tetanus originate in our hospital patients in seven years. The usual dose given is 1,500 units.

We give the serum therapeutically when occasion arises. I have had occasion to see in a distant city a case of tetanus in which the symptoms developed nineteen days after the primary infection. Repeated injections of the serum were given and the more violent symptoms subsided. The patient had no muscular twitching for ninety-six hours before his death.

Bacterial vaccines have been administered by us in all types and classes of cases such as are met in an extensive surgical practice.

Beginning at the lesions of the skin, we have treated cases of acne, furunculosis and carbunculosis, and infections of the lymphatic glands. Next we shall consider the lesions of the respiratory tract, as chronic rhinitis, asthma, hay-fever, bronchitis, empyema, and tuberculosis with mixed infection. Next chronic suppurations, including osteomyelitis and infected surgical wounds of all descriptions. Following this, chronic enteritis and kidney and bladder infections not associated with obstruction by foreign bodies. Then lowgrade pyogenic or saprophytic infections, and last, but most extensive, has been our experience with vaccines in all types of acute and chronic joint infections.

Bacteriologically our cases have included almost every variety of bacteria with varying degrees of virulence. Most common among them we found the Staphylococcus aureus and albus, the streptococcus, pneumococcus, the Bacillus coli, and the Bacillus pyocyaneus.

Detailed Considerations. The principles underlying the successful treatment of acne vulgaris

are:

1. The determination and isolation of the causative organisms.

2. The knowledge and experience necessary for the determination of the dosage and the interval between inoculations.

3. The general condition of patient and the degree of virulence of the infective organism. In the series of 15 cases treated by us we found that the Staphylococcus albus was present in 90 per cent of the cases.

Theoretically, the acne bacillus should be present in every case. The difficulty of growing this bacillus is the principal obstacle to the successful treatment of acne vulgaris. This organism requires from five to ten days on the most suitable

culture media, one of which is the 1 per cent oleic acid agar.

Another reason for failure is the fact that the dosage of the acne bacillus vaccine has been much too large. In all these cases it is also necessary to watch and modify the coagulability of the blood. If this is too rapid, the patient should have large doses of sodium citrate; on the other hand, if the coagulability is too slow and the patients do not improve immediately, it is necessary to give a number of 10 grain doses of the calcium lactate.

We do not encourage this type of cases, but many of them came to us for some surgical lesions. Several of them had made "the rounds," and finally appealed to us for the vaccine treatment.

In two of our cases all the lesions subsided after three or four injections, and have not returned. Some of the cases improved so rapidly after the fifth or sixth injection. One patient was not improved at all.

The furuncles and carbuncles, of which we have treated 20 cases, have all made rapid and complete recovery. The autogenous vaccines should always be given if one wishes to obtain the best results. One of our cases, who had previously been treated with mixed stock vaccines over a period of seven months, responded immediately after three injections of autogenous vaccines. Eighty per cent of these cases showed the Staphylococcus aureus.

Glandular infections, not tubercular, are due to the Staphylococcus aureus and albus, occasionally combined with the pneumococcus. Twentyfive cases of this type treated with auto-vaccines showed rapid improvement, and the majority of them went right on to ultimate complete recov

ery.

We have had one case of chronic rhinitis in a doctor, the most prominent symptom of which was a very annoying sneezing, which was treated with a mixed vaccine which contained the streptococcus and pneumococcus. The sneezing very materially improved after the fourth or fifth injection.

We have treated six cases of chronic asthma in which we were able to demonstrate the pneumococcus combined in the other five. In one we noted an aggravation of the symptoms from ten to eighteen hours after the injection, but a

marked improvement after the third dose. The other cases, one of which was not improved materially, did not show the reaction as did the first case. Another case of asthma, treated at the hospital under our direction, showed no pathogenic bacteria in any of the bronchial sections that were obtainable. We advised the use of a mixed pneumo- and staphylococcus vaccine. Later the interne informed me the patient's condition was greatly improved and he has had no more attacks.

Last August we treated three cases of hayfever with a mixed stock vaccine containing for the most part the streptococcus. Two of these cases did not find it necessary to change climate during the hay-fever season, as had been their custom for many years.

Much has been written in recent months on the use of vaccines in pertussis. Zahorsky has reported 40 cases treated with favorable results in some cases. In not a single case in which the vaccine was used did any secondary infectious process appear. His best results were obtained when the vaccine was given at the height of the disease.

Graham who reported 24 cases, believes that the course of the disease is shortened and improvement almost always continued after the injections were discontinued. He observed benefit in 17 cases. Ladd reported a series of cases. After the injections mothers stated that severity and number of paroxysms were diminished.

Four cases of purulent empyema with drainage were treated by us with auto-vaccines, resulting in a rapid diminution of the amount and odor of the discharge and ultimate permanent healing. Two patients had a green pus discharge, which delayed improvement for some weeks until a suitable autogenous vaccine of the Bacillus pyocyaneus could be prepared.

In tuberculosis with mixed infection we have made it a rule to use the vaccine, together with the tuberculin injections. The vaccines are given every four or five days, and the tuberculin once each week. Both injections should never be made on the same day, as that would interfere with the temperature reaction, which is the most important guide to the size of the tuberculin dosage.

Acute and chronic surgical infections of all types offer the largest field for vaccine therapy. Under this we include all cases which are pri

marily septic, incised or gunshot wounds, and osteomyelitis not associated with sequestra and not containing large cavities, pelvic infections, suppurations, appendicitis cases, etc. In a series of over 50 cases of this type we used the autogenous vaccines in about 90 per cent and the mixed vaccines in about 10 per cent. These cases, second to the pyocyaneus cases, give the most brilliant results. If the vaccines are properly made and the dosage and intervals properly graduated, this type should yield cures almost uniformly. The one reason for failure is the fact that the vaccines are not changed with sufficient frequency. A new vaccine should be made about every two weeks, to make sure that the organisms doing the most harm are incorporated in the vaccine. No culture can be transplanted more than two or three times. without detrimental attenuation.

Another large field for biologic therapy is the kidney and bladder infections not associated with foreign-body obstruction or organic obstructions. The colon bacilli, staphylococci and streptococci, and often the penumococci gain entrance to the urinary tract and cause marked frequency and often painful micturition. One of the most striking cases of this type came to the hospital late in October, 1911, suffering from frequent and painful urination, extending over a period of about ten months. After taking an accurate clinical history, it was found that this patient had suffered from dysentery of several weeks' duration, and that her trouble dated from that time and became progressively worse. Cystoscopic examination showed no pus coming from either ureter, and the absence of a neoplasm or foreign body in the bladder. The culture from the urine made the diagnosis of cystitis due to the Bacilli coli and staphylococci. The auto-vaccines were given about every five days, and the patient made a rapid and complete recovery and has remained permanently well.

Another case, Mr. D., came to the office on December 11, 1911, giving the history of a continued illness for four months, with repeated daily attacks of severe pain deep in the perineum, associated with chills and high temperature, frequent urination (every quarter to one hour), and large quantities of pus in the urine. Prostate gland not perceptibly diseased. Culture of the urine showed Bacilli coli and staphylococci. Vaccines were made and the patient improved rapidly for

several weeks, followed by a period of non-improvement for a few weeks. Another culture showed that all the staphylococci had disappeared, leaving a pure culture of Bacillus coli, from which a vaccine was made and injected in the usual way. This was in February, 1912. On April 1, 1912, Dr. Murphy made the following note: "The pains are all gone. Only one morning urination. The urine is absolutely normal." [Case has been permanently cured. July 2, 1913, patient reports himself perfectly well, with no evidence of return.]

Another patient, Mrs. G., suffering from a chronic cystitis, showed a bacillus not unlike the Bacillus pyocyaneus. The stock pyocyaneus vaccines were given at four-day intervals, and in a short time the pus-cells decreased and those that were present were greatly fragmented. This patient made a complete recovery, but had a recurrence from which she also recovered.

We have treated two cases of chronic enteritis, one of which was of tuberculin origin. This patient had from 10 to 20 bowel movements daily, most of them containing blood. The auto-vaccine was prepared, and after the first injection the blood ceased and the number of movements became less. The other case responded to none of the many vaccines given, and was later found to be a gas-bacillus infection.

We recommend the use of the prophylactic injections of the antityphoid vaccine in adults, especially in doctors, nurses and all persons liable to be exposed to the disease.

Three injections are given with an interval of 7-10 days between doses. The first dose contains 500 million bacteria and the second and third doses contain one billion bacteria each.

F. F. Russell, in the Jour. A. M. A., presented statistics on the antityphoid vaccinations in children between the ages of 2-16 years.

The dosage is based on body weight rather than the age of the child. About 359 children have been vaccinated in various parts of the United States. No harmful effects were noted and not one of the children has contracted the disease, though some were vaccinated three years ago. In the absence of final information as to the duration of immunity, now children are revaccinated after about three years.

In the last three years we have devoted our best efforts to the vaccine treatment of acute and